Zeitschrift für Immunitätsforschung, · Zeitschrift für Immunitätsforschung, experimentelle und...

Zeitschrift für Immunitätsforschung, experimentelle und klinische Immunologie

Herausgeber

H. BRANDIS, Bonn • H . DEICHER, Hannover • A. DE WECK, Bern K. O. ROTHER, Heidelberg • F. SCHEIFFARTH, Erlangen G. F. SPRINGER, Evanston, USA • C. STEFFEN, Wien

Mithe rausgebe r

I I . BEGEMANN, München • K. FED ERLIN, Ulm • H. FINGER, Krefeld K. FISCHER, Haniburg • A. HÄSSIG, Bern • K. HUMMEL, Freiburg W . KÖHLER. Jena • S. KOLLER, Mainz • M. KRÜPE, Fulda K. LENNERT, Kiel • E. LETTERER, Tübingen • E. MACHER, Münster H. J. MÜLLER-EBERHARD, La Jolla, USA • W. MÜLLER-RUCHHOLTZ, Kiel E. F. PFEIFFER, Ulm • 0. PROKOP, Berlin • W. RAPP, Heidelberg G. RIETHMÜLLER, Tübingen • H. SCHMIDT J, Bern • H. G. SCHWICK, Marburg W. SPIELMANN, Frankfurt * G. UHLENBROCK, Köln • H. WARNATZ, Erlangen M. WERNER, Pinneberg

148. Band

M i t 87 A b b i l d u n g e n

GUSTAV FISCHER VERLAG • STUTTGART • 1974/75

Inhaltsverzeichnis

I . Verzeichnis der in Band 148 enthaltenen Arbeiten

A B U N . R. J., and G. E . P I E P E R : Naturally Occurring An t ibody to H u m a n T11 v i 1 loo y tcs in No rmal Hab bit Scrum 208

A L B E R T , E. I ) . : The Significance of H L - A Disease Associations. . . . 382

A L B E R T , E . D. S. G R O S S E - W I L D E , H . A L B E R T , E . D. S. N E T Z E L , B . A L B E R T , K . D . S. R I T T N E R , C H . A L B E R T . E . Ü . S. S C H O L Z , S. A L B E R T . E . D. , S. S C H O L Z . J . B E R

T R A M S . R. XV. E W A L D , E . W E S T -I ' H A L . K . - W . H A T S C H K O , XV. S P I E L MANN and S. S E I D L : Representative H L - A Phenotype and Haplotype Frequencies of the German Populat i on 367

A N D R L I K O V Ä , J . S. W A R N K R , V .

B A L D O , B . A. , G . U H L E N B R U C H and B . S A L F N E R : Studies on. the Specificities of Various A n t i - H Reagents

B A N D L O W , G., U . K O S Z I N O W S K I und R . T H O M S S K N : Beeinflussung der 11 n m u n oge nit ä t von Mäuseleber-zellen durch das Mäusehepa t i t i s -virus (MHY-3) . Enhanced I m m u -nogenicity of Murine Liver Cells Infected by M H Y - 3

VAN B E K K l ' M , D . W . S. SfHAKKKK, U . W .

M K U T R A M S , J . S. A L B E R T , E. I ) . B E R T R A M S . «). s. G R O S S E - W I L D E , H . B E R T R A M S , J . , K . K U W E R T ,

I L C J R O S S E - W I I . D E , B . X E T Z E L and W . M E M P E L : Discasc-Susceptibility-Genotvpc for Mult ip le Sclerosis: SD-HL-A3-7 -LD-P i (7a )? . . . .

B E R T R A M S , J . , M . R O M M E L F A N G E R , M . S P I T Z N A S and E, K U W E R T : H L -A 2 7 and Acute Anterior Uve i t i s .

B E R T R A M S , J . , ( ) . T H R A E N H A R T , U . F E L D M A N S " and E . K U W E R T : H L - A Antigens in Cancel- of the Breast and the Collum uter i

B E R T R A M S , J . , O . T H R A E N H A R T , W . L U B O L D T und E . K L ' W E R T : His toid o in p a t i b i 1 i t ä t s - ( H L - A -) A n t i gen e bei 1000 Blutspendern. E in Bei t rag zum Aufbau einer Thrombozyten- , I jcu kozy1 en- und Knochenmark -Spenderbank

330

384

389

391

B E Y E R , J . S. Z A P A T A , M . B L O O , J . H . s. H E E K E R S , P. B O C K H O R N , H . S. S E I D L E R , D . B O R S T - L I L E R S , E . S. S Y B E S M A , J . P H . B O S M A N S K Y , K . S. R O V E N S K Y , J . B R A U N S T E I N E R , H . S. S C H W A R Z , S. B R E L I N S K A - P E C Z A L S K A , R .

s. M A C K I E W I C Z , S. B Ü L T M A N N , B . , H . F I N G E R , B . H E Y -

M E R , W . S C I i A C H E N M A Y R, H . H O F and ( ) . H A F E R K A M P : Adjuvancy of Streptococcal Nucleic Acids . . . 42

C A P P U C I N E L L I , P. s. R E I S S - G U T F R E U N D , R. J .

C A V A L L O , G . s. R E I S S - G U T F R E U N D , R . J .

C E B E C A U E R , L . S. R O V E N S K Y , J . C E B E C A U E R , L . S. S V E J C A R , J . C E L I N S K A - Sz P Y T K O , E .

s. M A C K I E W I C Z , S. C I N A D E R , B . : Six Years of the Inter

national Un ion of Immunological Societies, Presidential Report , Br ighton 1974 . 18

C O H E N , 11. s. E S K E N A Z Y , M .

D I C K E , K . A . s. S C H A E F E R , U . W . D O R K E N , B . S. L E N H A R D , V . D O M I N C I E , G . , and K. J O H N S O N : Iso

lat ion of Subcellular Fractions Containing Immunogenic Enterobacterial Common Antigen L

E B A T A , H . S. L I E , T . S. E L I Z A N , T . S. S. S C H W A R Z , J . E S K E N A Z Y , M . , and R. C O H E N : Sensi

tized and Stabilized Red Blood Cells Coated w i th V i A ntigcTi as a Sensitive Tool for the Detection of V i Agglut inins 264

E W A L D , R . W . S. A L B E R T , E . D .

F E D E R L I N , K . s. H E L M K E , K . F E L O M A N N , U . S. B E R T R A M S , J . F I N G E R , H . S. B Ü L T M A N N , B. F R I E D R I C H , H . , ( ) . S A M L A N D , J . K R Ü

G E R , W . G R O S S , U . H Ä C K E L L , K . K U N Z E U. C H . M Ü L L E R - E C K H A R D T : H L - A - A n t i g e n e bei Myasthenia gravis 394

http://Wii.de

468 • I nha l t sve r ze i chn i s

G A B L , F . s. S C H W A R Z , S . G Ö T Z E , D . , and R. A . R E I S F E L D : I m

mune Response to Soluble H - 2 K D

Antigens 4 5 G R O S S , W . s. F R I E D R I C H , H . G R O S S E - W I L D E , H . s. B E R T R A M S , J . G R O S S E - W I L D E , H . S. N E T Z E L , B . G R O S S E - W I L D E , H . , W R . M E M P E L , B .

N E T Z E L , E . D . A L B E R T , S. S C H O L Z , W . L U B O L D T , E . K U W E R T and J . B E R T R A M S : Def ini t ion and Genetics of L D Specificities in a German Populat ion and their Relat ion to the H L - A System 3 7 6

D E H A A N , M . A . M . s. S Y B E S M A , J . P H . H Ä C K E L L , U . S. F R I E D R I C H , H . H Ä N S C H , G. S. R O T H E R , U . H A F E R K A M P , 0 . S. B Ü L T M A N N , B . H A R M S , K . S. S C H O L Z , S . H E D S T R Ö M , K . - G . S. P R A S A D , C. B . H E I D E , K . - G . S. H U M M E L , K . H E L M K E , K . , U . W Ö L K E und K . F E -

D E R L IN : Eine immunfluoreszenz-optische Methode zum Nachweis von A n t i - D N S - A n t i k ö r p e r n . . . 162

H E R R L I N G E R , J . D . , and W R . M Ü L L E R -R U C H H O L T Z : I n h i b i t i o n of A n t i body Product ion of Sensitized Animals . I I I . Effects of a Combined Azathioprine/Antigen Treatment . 2 3 5

H E Y M E R , B . s. B Ü L T M A N N , B . H O F , H . S. B Ü L T M A N N , B . V A N H O O F F , J . P., and H . M . A . S C H I P

P E R S : The Influence of H L - A Matching, ABO Bloodgroup System, Transport i n Euro transplant and the Results of the H i g h Urgency List '. 3 9 6

H U B E R , H . s. S C H W A R Z , S. H U M M E L , K . , M . V A M O S I und K . - G .

H E I D E : Berechnung der Vater -Schaftswahrscheinlichkeit bei b lut gruppenserologischer Begutach -tung der Abs tammung von Z w i l l i n gen und Geschwistern. I I I . M i t t e i lung: Praktische Erfahrungen mi t der Methode an einer g röße ren Zahl von Zwill ings- u . Geschwisterfä l len 2 9 9

J O H N S O N , E . S. D O M I N G U E , G. J Ö K A Y , I . , and E . K A R C Z A G : Cellular -

and Permeability-Changes in the Peritoneal Cavi ty of Mice after I n jection of Endo tox in . - Zellular -und P e r m e a b i l i t ä t s v e r ä n d e r u n g e n i n der B a u c h h ö h l e von M ä u s e n nach Endotoxin in jek t ion 4 1 4

J U N G , M . S. K R E C H , U .

K A A R S S I J P E S T E Y N , J . A . s. S Y B E S M A , J . P H .

K A R C Z A G , E . S. J Ö K A Y , I . K N A P P , W . : Interactions of the T h i r d

Component of Complement (C3) w i t h Cross-Linked Dext ran . I . B i n d ing of C3 f rom Norma l H u m a n Serum to Cross-linked Dext ran as Defined by Quant i ta t ive I m m u n o fluorescence 213

K N A P P , W r., J . M E N Z E L and C. S T E F F E N : Microfluorometric Evalua t ion of Anti-Collagen-Antibodies w i t h the Defined Ant igen Substrate Spheres (DASS) System 132

K O S Z I N O W S K I , U . S. B A N D L O W , G . K O S Z I N O W S K I , U . , B . V O L K M A N N and

R. T H O M S S E N : I n v i t r o Demonstrat i o n of Cell-Mediated I m m u n i t y to Vaccinia Virus i n Man 451

K R E C H , U . , M . J U N G , P. C. P R I C E , G . T H I E L , D . S E G E and F . R E U T T E R : Virus Infections in Renal Transplant Recipients 341

K R Ü G E R , J . s. F R I E D R I C H , H . K U N S T , V . A . J . M . s. R E E K E R S , P. K U N Z E , K . S. F R I E D R I C H , H . K U W E R T , E . S. B E R T R A M S , J . K U W E R T , E . S. G R O S S E - W I L D E , H .

L A U C H A R T , W . S. S E I D L E R , D . L E N H A R D , V . , B . D Ö R K E N and M . A .

L E O N : Model of a modified Mixed Lymphocy te Culture -- Acceleration and Ampl i f ica t ion of the Reaction by Pretreatrnent w i t h Con A and Anti-Con A 404

L E O N , M . A . s. L E N H A R D , V . L I E , T . S., H . N A K A N O , H . E B A T A

und B . M O R I T Z : Akt ives Enhancement von Hundeleberallotransplan-ta ten 62

L O R E N Z , H . S. R I T T N E R , C H . L U B O L D T , W r. s. B E R T R A M S , J . L U B O L D T , W . S. G R O S S E - W I L D E , H .

M A C K I E W I C Z , S., R. B R E L I N S K A -P E C Z A L S K A and E . C E L I N S K A -S Z P Y T K O : Cytochemical Studies of Rosette-Forming Cells 462

M A R C O L O N G O , R., and C. M A R S I L I : Serum I g D and I g E in Rheumatoid A r t h r i t i s 285

M A R S I L I , C. S. M A R C O L O N G O , R . M A R T H Y , H . - J . : Nachweis und Bedeu

tung einer h ä m a g g l u t i n i e r e n d e n Substanz aus der H a u t von Cephalo-poden 225

M A Y R , W . R . : Molekulare Assoziation des ß. 2-Mikroglobiilins m i t serologisch definierbaren Merkmalen an der L y m p h o z y t e n o b e r f i ä e h e . . . 92

M C S H I N E , R. L . S. R E E K E R S , P. M E M P E L , W . S. B E R T R A M S , J .

Inha l t sve rze i chn i s 469

MKMIM-IL, W . S. G R O S S E - W I L D E , H . M E M P E L , W . S. X E T Z E L , B . M E N Z E L . J. S. K N A P P , W . M E N Z E L , J . S. K O T H E R , U . M Ö K I T Z , B . s. L I E , T . 8 . M UELLER -ECKHARDT, C H .

s. F R I E D R I C H , H . M 1J L L E R - I i UCHHOLTZ, W .

s. H E R R L I N G E R , J . 1 ) . M Ü L L E R - K I C H H O L T Z , W .

s. R A T S C H K O , K . - W . M i ' L L E R - R r c H i i o L T Z , W .

s. W E S T P H A L , E .

X AK A NO , H . s. L I E , T . S . X E T Z E L , B . S. B E R T R A M S , J . X E T Z E L , B . S. G R O S S E - W I L D E , H . X E T Z E L . B. , H . G R O S S E - W I L D E , E . D .

A L B E R T and W . M E M P E L : L D T y p ing w i t h Defined L D Heterozygous Reference Cells " . . . 3 7 9

P Ä i . E K , V . s. W A G N E R , V . P A P P A S . A . , and G . S C H W A R Z E : I n v i t ro

Dei nonstrat ion of Cell-Med iated I m m u n i t y against H u m a n Renal Carcinoma Assessed by the Leukocyte Migra t ion Assay 1.42

P A S T N E R , D . S. S C H W A R Z , S . P E K Ä R E K , J . S. R O V E N S K Y , J . P E K Ä R E K , J . S. S V E J C A R , J . P I E P E R , G . K S. A B L I N , R. J . P R A S A D , C. B. , and K . - G . H E D S T R Ö M :

Study on Exper imental Vaccinia Virus Infect ion in Mice w i t h Special Reference to Host Defence Mechanisms 97

P R I C E , P. C. s. K R E C H , U .

R A T S C H K O , K . - W . S. A L B E R T , E . D . R A T S C H K O , K . - W . , H . S T A M M ,

L. W E S T P H A L and \V. M Ü L L E R I N I C H HO L T Z : Search for H L - A A n t i genic Determinants in Bacteria and Rodents 4 0 7

R A T S C H K O , K . - W . s. W E S T P H A L , E. R E E K E R S , P., R . L . M C S H I N E , J . H .

B L O O and V. A . J . M . K U N S T : Posi t ive Indirect Ant ig lobu l in Test Caused by Leucocyte Antibodies . 4 0 2

R E D A , I . , G . WlTTM A N N Ulld R . SivODA : Der X a c h weis a n t i k ö r p e r b i l d e n d e r Zellen mit Hi l fe der lokalen H ä m o -lyse im Gel ( L H G ) bei Mäusen und Sch we i neu nach I m munisierung mit infekt iösein oder inakt iv ier tem Maul- und Klauenseuchevirus . . 119

R E I S F E L D , R . A. S. G Ö T Z E , D . R E I S S - G U T F R E U N D , R . J . , P. C A P P U C I -

N E L L I and G. C A V A L L O : The Soluble Antigens of Ricket ts ia prowazeki, R. t y p h i and R . canada. Investiga

tion, of their Interrelat ionship by Various Serological Methods . . . 3 1 5

R E N W R A N T Z , L . , Ulld G. UlILENBRUCK : B l u t g r u p p e n ä h n l i c h e Substanzen in einigen marinen 1 nvertebraten. I I . E i n durch Laktose hemmbares Agglu t in in neben einem Blu tgruppen-A-ähn l i chen Glykoprotein im Serum des gemeinen Kraken Octopus vulgaris (Lam.) 16

R E U T T E R , F . s. K R E C H , U . R I T T N E R , B . S. R I T T N E R , C H . R I T T N E R , C H . , B . R I T T N E R , S. S C H O L Z ,

H . L O R E N Z and E. D . A L B E R T : Data on a «Xew» Linkage Group: H L - A -B f (Factor B of t heProperdin System) 381

R I T T N E R , C H . S. W A I Y A W U T H , V . R O M M E L F A N G E R , M . S. B E R T R A M S , J . R O T H E R , K . S. R O T H E R , Ü . R O T H E R , U . , G. H Ä N S C H , J . M E N Z E L

and K . R O T H E R : Deviated Lysis : Transfer of Complement L y t i c Act i v i t y to Unsensitized Cells. I . Generation of the Transferable A c t i v i t y on the Surface of Complement Resistant Bacteria 172

R O V E N S K Y , J . s. S V E J C A R , J . R O V E N S K Y , J . , J . S V E J C A R , L . C E B E

C A U E R and J . P E K Ä R E K : Studies of Delayed Hypersensi t ivi ty in Rheumato id A r t h r i t i s 3 9

R O V E N S K Y , J . , K . T R N A V S K Y , J . P E K Ä R E K , J . S V E J Ö A R and K . B O S M A N S K Y : Migrat ion Inh ib i t o ry Factor in Gold Hypersensi t ivi ty . 112

S A L F N E R , B . S. B A L D O , B . A. S A M L A N D , 0 . s. F R I E D R I C H , H . SC H A C H E N M A Y R , W . S. B (' L T M A NN , B. S C H A E F E R , U . W . , K . A . D I C K E , D. W .

VAN B E R K U M and C. G . S C H M I D T : Cryopreservation of Bone Marrow. 3 9 9

S C H A T T E N K I R C H N E R , M . S. S C H O L Z , S. S C H I E K , VV.: Untersuchung zur An

t i k ö r p e r b i l d u n g in v i t ro . 111. In dukt ion einer p r i m ä r e n und sekundä ren Immunan twor t i n v i t ro . Studies on the in v i t ro formation of antibodies. I I I . Induct ion of pr imary and secondary immune response in v i t ro 431

S C H I P P E R S , H . M . A . s. V A N H O O F F , J . P .

S C H M I D T , C. G. S. S C H A E F E R , U . W T . S C H M I T T , H . S. Z A P A T A , M . S C H O L Z , S. S. A L B E R T , E . D . S C H O L Z , S. S. G R O S S E - W I L D E , H . S C H O L Z , S. S. R I T T N E R , C H . S C H O L Z , S., M . S C H A T T E N K I R C H N E R ,

K . H A R M S , I . S T E I N B A U E R - R O S E N -T H A L and E . D . A L B E R T : Fami ly St udies in H L - A Associated Diseases 3 8 7

470 • Inha l t sve rze i chn i s

S C H W A R T Z , J . , and T . S. E L I Z A N : Altered Immunologic Specificity of Cells Infected w i t h Herpes Simplex Vi rus : Recognition by H u m a n A n tiserum 291

S C H W A R Z , S., F . G A B L , D . P A S T N E R , H . H U B E R und H . B R A U N S T E I N E R : Immung lobu l in E bei chronischer lymphatischer L e u k ä m i e und monoklonalen Gammopathien . . . . 196

S C H W A R Z E , G. S. P A P P A S , A . S E G E , D . S. K R E C H , U . S E I D L , S. S. A L B E R T , E . D . S E I D L E R , D . , G. T R A U T W E I N ,

W . L A U C H A R T und H . B O C K H O R N : Morphologische Befunde an der Leber des Schweines nach allogener orthotoper bzw. aux i l i ä re r hetero-troper Transplantat ion I

Six th Annual Meeting of the German Tissue T y p i n g Laboratories . . . . 367

S K O D A , R. S. R E D A , I . S P I E L M A N N , W . S. A L B E R T , E . D . S P I T Z N A S , M . S. B E R T R A M S , J . S T A M M , H . S. R A T S C H K O , K . - W . S T E F F E N , C . S. K N A P P , W . S T E I N B A U E R - R O S E N T H A L , I .

s. S C H O L Z , S. S Y B E S M A , J . P H . , M . A . M . D E H A A N ,

E. B O R S T - E I L E R S and J . A . K A A R S S I J P E S T E Y N : Demonstrat ion of A n tibodies in Potent ial K i d n e y Reci-pientsby Meansof aLong Incubat ion Lymphocy to tox i c i t y Cross-Match . 397

SvEJCAR, J. , J . R O V E N S K Y , J . P E K Ä R E K , D . Z I T N A N and L . C E B E C A U E R : Delayed Hypersens i t i v i t y to D N A i n Systemic Lupus Erythematosus and Related Diseases 244

S V E J C A R , J . S. R O V E N S K Y , J .

T H I E L , G. S. K R E C H , U . T H O M S S E N , R. S. B A N D L O W , G. T H O M S S E N , R. S. K O S Z I N O W S K I , IT. T H R A E N H A R T , O . S. B E R T R A M S , J . T R A U T W E I N , G. S. S E I D L E R , D . T R N A V S K Y , K . S. R O V E N S K Y , J .

U H L E N B R U C K , G. S. B A L D O , B . A . U H L E N B R O C K , G. S. R E N W R A N T Z , L .

V A M O S I , M . S. H U M M E L , K . V O L K M A N N , B . S. K O S Z I N O W S K I , U .

W A O N E R , V . , J . A N D R L I K O V Ä and V . P A L E K : Immunoglobul ins under the Influence of Nonspecific Factors. I I . The Influence of Work-stress on Levels of Immunoglobul ins ( IgG, Ig A, I g M ) of Miners i n Uran ium Mines 356

W A H L , R., et I . W A U Q I E R : Phagozytose de Streptocoques Hemolyt iques du Groupe A par des Macrophages de Souris. Influences Respective* LMnfection Prealable des Animaux et du Degre de Virulence de la Souche 273

W A I Y A W U T H , V . , and C H . R I T T N E R : Improved Technique for H L - A T y p i n g i n Dr ied Blood Stains: Modified Ex t rac t ion of Stains and Appl ica t ion of the Microcomple-ment fixation 404

W A U Q U I E R , I . s. W A H L , R . W E I L A N D , E . , und F . W E I L A N D :

Transplantierbarer Aszitestumoraus einer Maus mi t einem Moloneysark o m : Tumor-, Antigen- und Viruseigenschaften 151.

W E I L A N D , F . s. W E I L A N D , E . W E S T P H A L , E . S. R A T S C H K O , K . - W . W E S T P H A L , E . S. A L B E R T , E . D . W E S T P H A L , E . , K . - W . R A T S C H K O and

W . M Ü L L E R - R U C H H O L T Z : New Evidence for Newer H L - A Specificities 371

W I T T M A N N , G . S. R E D A , 1. W Ö L K E , U . S. H E L M K E , K .

Z A P A T A , M . , H . S C H M I T T and J . B E Y E R : The Le a -Ant igen on H u m a n L y m phocytes 410

Z I T N A N , D . S. SVE.TCAR , J .

Buchbesprechung • Book Review

W E I L E R , E . : I m m u n i t ä t s f o r s c h u n g u. das Dogma der molekularen Biologie. Bespr. von H . F I N G E R . . . . 90

Sachverzeichnis 471

I I . Sachverzeichnis

Aalserum (anti-H) 330 AHO bloodgroup system,

Euro transplant 396 AdjuvansWirkung, von

S t r e p t o k o k k e n - N u k l e i n s ä u r e n . . 425 Agglu t in in bei Invertebraten . . . . 16 Agglu t in in , V i - , Nachweis 264 AI logene Transplantat ion,

morphologische Befunde 1 Allotransplantate, Hundeleber,

aktives Enhancement 62 Alternate pathway of complement

activation 213 Ant i-DNS-Ant ikorper, i m m u n -

fluoreszenzoptische Methode . . . 162 Ant igen, gemeinsames

bei Enterobacteriaceae 23 A nt ige I i , H istokoi npat ibi 1 i t ä t s -

siehe H L - A Antigen, Le a - , an Lymphozyten . . 410 Antigene Determinanten, H L - A -,

bei Bakterien und Nagetieren . . 407 Antigene, H-2 K c l , lösl iche,

Immunantwort gegen 4") Antigene, H L - A - , bei Blutspendern . 73 Ant igene, lösliche,

bei R. prowazeki, R. t y p h i und R. canada 315

A111 igensu bst ratsystem, definiert es (DASS) zum Nachweis von A n t i Ko l l agen-Ant ikö rpe rn 132

Ant ig lobu 1 intest, positiver indirekter, durch Leukozyten-Ant ikörpe r 402

Anti-H-Substanzen, Spezifi tät . . . 330 A11t i l i u m an t h y m ozy t englobu l i n ,

vom Kaninchen 208 A n t i k ö r pe r, A n t i - D N S -,

immunfluoreszenzopt ische Methode 162 Ant ikörper , bei vorgesehenen

Empfänge rn für eine Niere . . . . 397 An t ikö rpe r gegen Kollagen,

mikrofluorometrische Bestimmung 132 A n t i k ö r p e r gegen Vi ren ,

nach Nierentransplantation . . . 341 Ant ikörper , Leukozyten- 402 Ant ikörpe r , na tü r l i ch vorkommende,

gegen m en sc h lie h e T h y m o zy ten in Kaninchen-Seren 208

Aii t iköi 'perbi ldende Zellen, Nachweis m i t Jen le - PI aqu e - Tec h ni k, nach Immunisierung m i t MKS-Vi rus . . 119

An t ikö rpe r -B i ldung , Hemmung bei sensibilisierten Tieren 235

An t ikö rpe rb i ldung , in v i t r o . . . . 431

A nt i - Kollagen - A n t i kö r per, mikrofluorometrische Best immung 132

A r t h r i t i s , r heu m a t o i d e, I g D und I g E bei 285

A r t h r i t i s, r heu mat o i de, verzöger te Ü be re m pfi n d lieh ke i t bei 39

A-Streptokokken 273 Aszi test umor, t ransp 1 ant ierbarer,

von Maus m i t Moloneysarkom, Eigenschaften 151

Azathiopr in - A n t igen- Behand I ung, W i r k u n g auf immunologische Reaktionslage 235

Bakterien, HL-A-Determinanten. . . 407 b l u t g r u p p e n - ä h n l i c h e Substanzen,

bei Inver tebraten 16 blutgruppenakt ive (ant i -H)

Substanzen, Spezifi tät 330 bl u t gruppenser o 1 og i sc he

Begutachtung 299 Blutspender, Virusinfektionen bei . . 341

C3, Abb ind ung an quervernetztes Dext ran . . . . 213

C-Akt iv i t ä t , lytische, Ü b e r t r a g u n g (deviated lysis) 172

Carzinom, renales, Nachweis zellvermittelter I m m u n i t ä t . . . . 142

Cephalopoden, h ä m a g g l u t in.ieren.de Substanz aus der Haut 225

Collum uteri und H L - A 391 Cryopreservation, Knochenmark . . 399 Cytomegalic-Virus 341

DASS 132 definiertes Ant igensubstrat

System (DASS) 132 deviated lysis, Ü b e r t r a g u n g

lytischer C-Ak t iv i t ä t 172 Dextran , quervenletztes,

Abbindung von C3 213 DNS, ve rzöger te

Ü b e rem | )fi 11 d 1 ich ke i t gegen . . . . 244

Endotoxin , Wirkungen in der P e r i t o n e a l h ö h l e . 414

Enhancement, aktives, von Huiideleberal lotrai isplantatei i . 63

En terobac ter i aceae, gemeinsames Antigen 23

Faktor B 381 Gammopathien, monoklonale, I g E . 196

http://ieren.de

472 Sachverzeichnis

Goldüberempf ind l i chke i t , M I F bei . 112 H ä m a g g l u t i n a t i o n , passive zum

Nachweis von Vi-Agglut inin . . . . 264 h ä m a g g l u t inierende Substanz,

aus der Hau t von Cephalopoden . 225 H ä m a g g l u t i n i n bei Invertebraten . . 16" H ä m o d ialy se - Pat ienten,

Virusinfektionen bei 341 H ä m o l y s e im Gel, lokale, Nachweis

a n t i k ö r p e r b i l d e n d e r Zellen nach Immunisierung mi t MKS-Vi rus . . 119

Hap lo typ (HL-A)-Häuf igke i t . . . . 307 Hepat i t i s , bei Mäusen 254 Herpes simplex-Virus 341 Herpes s iDiplex-Vi rus ,

antigenetische v e r ä n d e r t e Zelloberfläche bei In fek t ion mi t 291

Hist ok o l n p at i b i 1 i t ä t s - An t igene siehe H L A

H i s t o k o m p a t i b i l i t ä t s - ( H L - A - ) Antigene bei 1000 Blutspendern . 73

H-2 K d -An t igene , Immunan twor t gegen lösliche . . 45

H L - A 2 7 und akute vordere Uvei t is . 389 H L - A Antigendeterminanten

bei Bakter ien und Nagetieren . . 407 HL-A-Ant igene bei Blutspendern . . 73 H L A , Assoziation bei Krankhei ten . 382 H L - A - assoz i ie r t e K r a n k h eiten,

Familienstudien 387 H L - A bei Carzinom der Brust

oder des Collums 391 H L - A - m a t c h i n g Eurotransplant . . 396 H L - A und Myasthenia gravis . . . . 394 H L - A - P h ä n o t y p , Häufigkei t . . . . 367 H L - A - P h ä not y p

bei mul t ip ler Sklerose 384 HL-A-Spez i f i t ä t en , neuere 371 H L - A -Ty pisier ung

mi t getrocknetem Blut 404 H L - A - B f , eine neue linkage Gruppe . 381 HL-A-Sys tem, Beziehung zu L D . . 376 H u n d e 1 e be r a 11 o t I - an s p 1 an tat e,

aktives Enhancement 62

l g A -Spiegel 356 I g D . . ' 285 IgE 196, 285 IgG-Spiegel 356 IgM-Spiegel 356 In imunant wor t ,

gegen lösliche H-2K c l -An t igene . . 45 In imunant wor t , I n d u k t i o n der . . . 431 immunfluoreszenzoptische Methode,

zum Nachweis von A n t i - D N S -A n t i k ö r p e r n 162

Immung lobu l in siehe auch I g Immung lobu l in ,

Einf luß unspezifischer Faktoren . 356 Immung lobu l in D

bei rheumatoider A r t h r i t i s . . . . 285

Immunglobu l in E. bei chronisch lymphatischer L e u k ä m i e und monoklonalen Gammopathien . . 196

Immunglobul in E, bei rheumatoider A r t h r i t i s . . . . 285

Immunis ierung mit MKS-Vi rus , Nachweis a n t i k ö r p e r b i l d e n d e r Zellen mi t der Jerne-Plaque-Technik 1 19

I m m u n i t ä t , zellvermittel te, gegen renales Carzinom 142

I in m un i t ä t , ze 11 ver m i 11 elt e, gegen Vaccinia-Viru« 451

II n l n im oge n i ta t , ge steigerte, von Leberzellen, infiziert mi t M H V - 3 254

I m inunological Soc iet ies, international Union, 6 years . . . 187

i i n i n u n o 1 og i s c h e S p e z i fi t ä t , v e r ä n d e r t e der Zelloberfläche bei In fek t ion mi t Herpes-simplex-Virus 291

Immunsuppression 235 Interferon 97 Intel-national Union of

Immunological Societies, 6 years . 187 Tnvertebraten, a n t i - H , aus,

Spezif i tät 330 Invertebraten, b l u t g r u p p e n ä h n l i e h e

Substanzen bei 16

«lerne - PI aquo -Tech n i k, Nach weis von a n t i k ö r p e r b i l d e n d e n Zellen nach Immunisierung mi t M K S - V i r u s . . 119

K an i n c h en - A n t i h u I n an -thymozytenglobul in 208

Knochentnark, eryopreservation . . 399 Knochenmarkspenderbank 73 Kollagen, A n t i k ö r p e r gegen,

mikrofluorometrische Best immung 132 Komplement -Akt iv ie rung ,

alternate pathway 213 Komplement -Akt iv ie rung , an

komplementresistenten Bakterien . 172 K () i n | )1 e l n e n t - A k t i v i t ä t , 1 y t i sc h e,

Ü b e r t r a g u n g (deviated lysis) . . . 172 Komplement , 3. Komponente,

1111 erakt ion In i t (juer\ 7ernctztein Dextran 213

Le a -Ant igen an Lymphozyten . . . 410 Li)-Sj)ezif i täten,

Definition und Genetik 376 L D-Typing 379 Lebertransplantation, allogene

ortl iotope bzw. heterotope, morphologische Befunde 1

Lekt ine (an t i -H) . Spezif i tät . . . . 330 L e u k ä m i e ,

chronisch lymphatische, I g E . . . 196

Sachverzeichnis 473

L e u k o z y t e n - A n t i k ö r p e r , positiver indirekter Ant ig lobu l in test . . . . 402

Lymphozy tenku l tu r , siehe mixed I.e. 404 Leukozyten -M igrat ions-Versuch,

Nac hwe is zellverm i t te l ter I m m u n i t ä t bei renalem Carzinom . 142

Leukozytenspenderbank 73 Loligo vulgaris 225 Lupus erythematosus, ve rzöger te

Überempf ind l i chke i t gegen DNS bei 244 Lymphozy ten , Le a -An t igen an . . . 410 Lymphozy ten ,

rosettenbi 1 dende Zeilen, cytochemische Untersuchungen . . 462

lysis, deviated, Ü b e r t r a g u n g lytischer C-Akt iv i t ä t 172

M ä u s e h e p a t i t i s v i r u s , M H V - 3 , Beeinflussung der I m m u n o g e n i t ä t von Mäuse leberze l len 254

Mäuseleberze l len 254 M a k r o g l o b u l i n ä m i e Waldenstrom . . 196 Makrophagen, Mäuse- , Phagozytose

von A-Streptokokken 273 Makrophagen-Migrations-Hemm-

faktor, bei Goldüberempf ind l ichke i t 112 Mammakarz inom und H L - A . . . . 391 Maul - und Klauenseuche-Virus,

Immunis ierung m i t , Nachweis von ant i kö rpe rb i l denden Zeilen m i t der Jerne-Technik 119

M H V - 3 254 M I F 244 M I F bei Goldüberempf ind l ichke i t . . 112 M igrat ions -Hemmfakt or

bei Go ldüberempf ind l i chke i t . . . 112 M igrat ions-Hemmfaktor ( M I F ) . . . 244 Migrat ions-Inhibi t ionsfaktor . . 119, 142 Mikrokomplement f ixa t ion 404 Mixed lymphocyte culture 376 Mixed lymphocyte culture,

modifizierte 404 M K S - V i r u s 119 M LC 376 Moloneysarkom, bei Mäusen ,

transplantierbarer Aszitestumor bei 151 Morbus Waldenstrom 196 M ul t iple Sklerose,

k i 'ankh e i t sd i spo n i er en der Ph äno t y p 384 Myasthenia gravis und H L - A . . . . 394 Mye lom, multiples 196 M y x o v i r e n 341

Nagetiere, HL-A-Dete rminan ten . . 407 Nierentransplantat ion, A n t i k ö r p e r

bei vorgesehenen E m p f ä n g e r n . . 397 N i erent rans pi ant at i on,

Virusinfekt ionen nach 341 N i e r e n t r a n s p l a n t a t ü b e r l e b e n s z e i t . . 396 N u k l e i n s ä u r e n , Streptokokken-,

Adjuvanswi rkung 425

Octopus vulgaris, b l u t g r u p p e n ä h n l i c h e Substanz . . 16

Phagozytose, von Streptokokken durch M ä u s e m a k r o p h a g e n . . . . 273

Picornaviren 341 Plasmozytom 196 Polyar thr i t i s , p r i m ä r chronische . . 244 Polyomaviren 341 Properdin-System 381

Respiratory syncytial Virus . . . . 341 rheumatoide A r t h r i t i s ,

I g D und I g E bei 285 Rheumatoide A r t h r i t i s ,

ve rzöger te Überempf ind l i chke i t . . 39 Ricketts ia canada,

lösliche Antigene bei 315 Ricket ts ia prowazeki,

lösliche Antigene bei 315 Ricket ts ia t y p h i ,

lösliche Antigene bei 315 Rosettenbildende Zellen,

zytochemische Studien 462

Sarkom virus Moloney 151 Schwein,

al 1 ogene Leber t r ansplaut at ion, morphologische Befunde 1

S D - H L - A 3 - 7 - L D - P i ( 7 a ) , Genotyp . 384 Sixth Annua l Meeting, German

Tissue T y p i n g Laboratories, 11.-12.10.1974 367

Sklerodermie 244 Streptokokken der Gruppe A ,

Phagozytose durch M ä u s e m a k r o p h a g e n . . . . 273

Streptokokken - N u klei n s ä u r e n , Adjuvanswirkung 425

Thrombozyten-Spenderbank . . . . 73 T h y m o zy te n, me n sch 1 i c h e,

A n t i kör per gegen, in Kaninchen-Seren 208

Tintenfisch 225 Tissue Typ ing , Abstracts 367 Transpl an tat e, H undelebe ral lo -,

aktives Enhancement von . . . . 63 Transplantat ion, allogene orthotope,

bzw. heterotope, morphologische Befunde 1

transplant ierbarer Aszitestumor . . 151 Tumor , Aszites-, transplantierbarer,

bei Moloney-Sarkom 151 T y p i n g . . ' 367

Überempf ind l i chke i t , Gold-, M I F bei 112 Überempf ind l i chke i t , ve rzöger te ,

bei rheumatoider A r t h r i t i s . . . . 39

474 • Sachverzeichnis

Übe rempf ind l i chke i t , ve rzöger te , gegen DNS, bei system. Lupus erythematosus 244

Uvei t i s , akute vordere, und H L - A 27 389

Vaccinia-Virus-Infekt ion bei Mäusen , Abwehrmeehanismen 97

Vaccinia-Vi ins, zellvermittelte I m m u n i t ä t gegen . 451

Varizellen -zoster-Virus 341 Va t er seh aft s w all r sc he i n 1 i c h ke i t ,

Berechnung 299 Vi -Agg lu t in in , Nachweis mi t

sensibilisierten und stabilisierten Ery throzy ten 2()4

Virus , Herpes simplex 291 V i r us -1 n f ek t i onen,

nach Nierentransplantat ion . . . 341

Zellfraktionen, von Entero-bacteriaceae, gemeinsames Antigen 23

Zelloberfläche, antigenetische ve rände r t e durch Herpes-lnfektion 291

Zellvermit tel te I m m u n i t ä t gegen. renales Carzinom, Nachweis mit Leukozyten-Migrations-Versuch . . 142

Zell vermit te l te I m m u n i t ä t gegen Vaccinia-Virus, In -v i t ro -Xachweis . 451

Z. Immun.-Forsch. Bei. 148, S. 451-461 (1975)

H y g i e n e - I n s t i t u t der U n i v e r s i t ä t G ö t t i n g e n

In-Vitro Demonstration of Cell-Mediated Immunity to Vaccinia Virus in Man

ULRICH KOSZINOWSKI, BERND VOLKMANN and REINER THOMSSEN

W i t h 5 F igures • Rece ived A u g u s t 23, 1974 • A c c e p t e d September 1, 1974

Abstract Cell m e d i a t e d i m m u n i t y t o vacc in i a v i r u s i n m a n was s t ud i ed b y l y m p h o c y t e

t r a n s f o r m a t i o n . V a c c i n i a an t i gen , p ropaga t ed on B H K - 2 1 a n d V e r o cells, cou ld be used successfully for i n - v i t r o t e s t i n g after p a r t i a l p u r i f i c a t i o n as w e l l as crude infect ious homogenates . V a c c i n i a an t igen p repa ra t ions were effective b o t h i n the in fec t ive a n d the i n a c t i v a t e d s ta te . I n a c t i v a t i o n was u sua l l y accompanied w i t h a ce r t a in loss o f s t i m u l a t i n g a c t i v i t y . D e v e l o p m e n t o f cell med i a t ed i m m u n e response i n - v i t r o after f irst v a c c i n a t i o n was inves t iga ted i n 17 adu l t s . V a c c i n i a v i ru s specific l y m p h o c y t e t r a n s f o r m a t i o n was seen i n the second week after vacc inat i o n in al l cases. F o l l o w i n g r e v a c c i n a t i o n no increase o f l y m p h o c y t e t r ans fo rmat i o n r a t i o cou ld be observed i n 11 persons s tud ied . A t the same t i m e the t i t e r s o f h u m o r a l an t ibod ies were e leva ted .

Introduction

Host response to viral infections include humoral and cellular immune mechanisms. Recovery from viral infections may be dependent on antibodies; antibody formation is of undoubted value in preventing disease (3). However, the development of delayed hypersensitivity to viral antigens is well recognized and at least in some viral diseases recovery from the primary infection may be largely because of cellular immune mechanisms (2). In vaccinia infection it was supposed from early studies of v. PIRQUET (17) that cellular immunity might be of essential importance. In experimental vaccinia infection, impairment of host cell-mediated responsiveness by use of immunosuppressive agents increased morbidity and mortality (11). In humans progressive vaccinia occurred in subjects with defects of cell-mediated immunity (8). Thus measurement of antibodies seems not to be sufficient enough for the evaluation of the immune status of a vaccinated person. Cell mediated immunity (CMI) to vaccinia virus has been successfully demonstrated in animals (7, 9, 15, 16, 19). In humans CMI to vaccinia has been tested by intracutaneous application of antigen (14) and also in-vitro by using different assays (5, 10, 12, 20, 2 1 , 24). In comparing the results difficulties may arise because the immune response of a vacci-

4 5 2 • U . K O S Z I N O W S K I , B . V O L K M A N N a n d R . T H O M S S E N

nated individual to vaccinia antigen in-vitro may to some extent depend on the host cell used for preparation of the in-vitro test antigen ( 1 ) . We therefore investigated the immune response of vaccinated persons to infectious and inactivated vaccinia virus antigen by using the lymphocyte-transformation test. Secondly we investigated the onset of cell-mediated reactions in-vitro following primary vaccination and the possible booster reaction of cellular immunity after revaccina-tion.

Material and Methods

S u b j e c t s

17 i n d i v i d u a l s , r a n g i n g i n age f r o m 17-28 years were s t u d i e d for i m m u n e react ions after a p r i m a r y v a c c i n a t i o n . These subjects were vo lun tee r s , w h o req u i r e d v a c c i n a t i o n i n order t o o b t a i n an i n t e r n a t i o n a l t r a v e l cer t i f ica te . These persons h a d n o t received v a c c i n a t i o n i n c h i l d h o o d because o f cutaneous or o ther illnesses undergone b y themselves or o the r members o f t h e i r fami l ies . E l e v e n i n d i v i d u a l s o f the i n s t i t u t e s ta f f were checked for ce l lu la r a n d serological i m m u n e react ions after r evacc ina t i on .

V i r u s e s a n d c e l l c u l t u r e s

T h e f o l l o w i n g s t ra ins o f viruses were used for p r e p a r a t i o n o f i n - v i t r o an t igens : V a c c i n i a v i r u s s t r a i n W R w i t h t i t e r s o f 10 6» 5 T C T D 5 0 / m l , Ves icu la r s t o m a t i t i s v i ru s ( V S V ) s t r a i n I n d i a n a (10* T C I D 5 0 / m l ) . T i t r a t i o n o f vacc in ia v i r u s a n d V S V was p e r f o r m e d on V e r o cells.

Cell cu l tu res were g r o w n a n d m a i n t a i n e d i n Eagle 's m i n i m u m essential m e d i u m supp lemen ted w i t h 1 5 % c a l f se rum, 100 u n i t s o f p e n i c i l l i n / m l a n d 1 0 0 / / g / m l s t r e p t o m y c i n . Viruses were p ropaga ted on p e r m a n e n t m o n k e y k i d n e y - e e lI cu l tu res (Vero) or on pe rmanen t b a b y hamste r k i d n e y - c e l l cu l tu res ( B H K - 2 1 ) .

- F o r v a c c i n a t i o n vacc in i a v i r u s s t r a i n Els t ree p f u 1 x 1 0 8 / m l ( L a n d c s i m p f a n -s t a l t D ü s s e l d o r f ) was used. I n t he case o f p r i m a r y v a c c i n a t i o n 1000 I U a n t i -v a c c i n i a - g l o b u l i n ( V a c c i n i a b u l i n ® / I m r n u n o ) was app 1 ied.

P r e p a r a t i o n o f a n t i g c n s

The infected cell cu l tures were d i s rup ted b y freezing a n d t h a w i n g 3 t imes af ter t i t r a t i o n , the cell homogenate was sonicated 3 x 1 0 sec w i t h a Branson Sonif ler a t stage 4. T h e m a t e r i a l was cen t r i fuged for 2 0 m i n u t e s at 2 5 0 0 r p m / 4 ° C t o r emove r o u g h cell f ragments . T h e supe rna tan t was subjected t o a pel let c e n t r i -f l i g a t i o n i n a B e c k m a n n cent r i fuge L 2 a t 4 ° C , 9 0 m i n u t e s 1 7 0 0 0 r p m in t he r o t o r S W 27 or t y p e 19 120 m inu t e s , 1 8 0 0 0 r p m . I n these steps the v o l u m e was reduced 100 : 1 . T h e pe l le t was suspended i n phosphate buffered saline pH 7.2 son ica ted aga in for 10 seconds and subsequent ly p u r i f i e d f r o m cell f ragments b y 5 m i n u tes 5 0 0 0 r p m 4 ° C c e n t r i f u g a t i o n i n an I E C - c e n t r i f u g e . T h e T C I D 5 0 / m l o f t h i s p r e p a r a t i o n reached usua l ly 1 X 10

8

—1 x 109

.

V i r u s i n a c t i v a t i o n

T h e in fec ted cell homogena te or p a r t l y p u r i f i e d m a t e r i a l was i n a c t i v a t e d w i t h f o r m a l i n 1 : 4 0 0 0 over 2 4 hours a t 3 7 ° C . A b u n d a n t f o r m a l i n was b o u n d w i t h s o d i u m pyrosu l f i t e i n e q u i m o l a r c o n c e n t r a t i o n . U V - i n a c t i v a t i o n was p e r f o r m e d b y i r r a d i a t i o n o f m i n u t e vo lumes i n a t h i n l aye r i n p e t r i dishes for 5 m i n u t e s a t a dis tance o f 10 c m f r o m t h e U V - s o u r c e ( o u t p u t 10 W a t t a t 2 5 4 n m ) .

Gel I -Media ted I m m u n i t y to V a c c i n i a V i r u s • 453

J j y I n p h o c y t e t r a n s f o r m a t i o n - t e s t

B l o o d was asept ica l ly r e m o v e d and m i x e d w i t h hepar in (25 U m l ) . Sediment a t i o n o f e r y t h r o c y t e s was pe r fo rmed af ter a d d i t i o n o f 1 m l 5 % D e x t r a n ( M G 250,000. Pharmac ia , Uppsa la ) t o 9 m l b lood and i n c u b a t i o n i n 4 5 ° i n c l i n a t e d tubes for 45 m i n u t e s at 3 7 ° C . The upper phase, r i c h in l y m p h o c y t e s was washed 2 t imes in H a n k s balanced salt s o l u t i o n , c o n t a i n i n g 2 % i n a c t i v a t e d fe ta l c a l f serum and 5 U h e p a r i n / m l a n d then suspended i n c u l t u r e m e d i u m ( T C M 199 w i t h 1 5 % i n a c t i v a t e d fetal ca l f se rum, 100 U p e n i c i l l i n and 100 / /g s t r e p t o m y cin) t o g ive a f inal concen t r a t i on o f 2 x 10G ce l l s /ml . Cul tu res were set u p i n plas t ic tubes (0 re ine r , N ü r t i n g e n , N o . 160S) c o n t a i n i n g 1 m l l y m p h o c y t e suspension a n d 0.2 m l o f a n t i g e n : v i r a l an t igen , non in fec t i vc tissue cell homogen ate or c o n t r o l an t igen o f V S V to w h i c h the p a t i e n t was not sensit ized. T h e a b i l i t y for unspecific t r a n s f o r m a t i o n was tested by a d d i t i o n o f P h y t o h e m a g g i u t i n i n (Wel lcome) 12.5 / / g / m l . E a c h test was run a t least 3 t imes . T h e tubes were i n cuba ted at 37 ° C for 5 days i n a h u m i d i f i e d 5 % C 0 2 a tmosphere . T w e l v e hours before the t e r m i n a t i o n o f cu l tu res 1.5//Ci 3 H - T h y m i d i n e (spec, a c t i v i t y 23 m C m m o l , Radiochemica ls A m e r s h a m , Buch le r , Braunschwe ig ) were added . T o terminate ' t he cu l tu res the tubes were cooled to 4 ° C , cen t r i fuged at 1000 r p m for 10 m i n u t e s , and the superna ta i i t s d iscarded. T h e sediments were suspended i n 10 m l phys io log ica l N a C l , f i l te red t h r o u g h m e m b r a n e filters, pore d iamete r 0 .2 / / ( S A R T O R I U S . G ö t t i n g e n ; Cat . N o 11307). and washed w i t h 1 0 m l ice co ld 5 % t r i ch lo roace t i c ac id ( T C A ) . T h e d r i e d filters were p u t i n t o v ia l s w i t h sc in t i l l a t i o n fluid a n d coun ted i n a L i q u i d s c i n t i l l a t i o n spec t rometer ( P a c k a r d Mode l 3380). T r a n s f o r m a t i o n r a t i o was ca lcu la ted as the r e l a t ionsh ip o f i n c o r p o r a t i o n o f r a d i o a c t i v i t y in acid insoluble D N A i n an t igen t r ans fo rmed cu l tu res t o c o n t r o l cu l tu res .

N e 11t r a I i z a t i o I i t e s t

Sera assayed for vacc in ia a n t i b o d y were heated a t 5 0 ° C for 30 m i n u t e s before assay and serially' d i l u t e d i n f o u r f o l d steps in Eagle s M E M . M i x t u r e s , c o n t a i n i n g 2 pa r t s o f d i l u t e d sera, 1 pa r t o f gu inea p i g complement (90 h e m o l y t i c U / m l , 1 : 15 d i l u t e d p r i o r to use) a n d 1 par t o f v i r u s suspension ( g i v i n g a final v i r u s concen t r a t i on o f 100 T C I D 5 0 / 0 . 2 m l o f m i x t u r e ) were i n c u b a t e d o v e r n i g h t a t 37 C. A l i q u o t s o f 0.2 m l / w e l l were t hen inocu la ted on mono laye r s o f V e r o cells, prepared in m i c r o l i t e r plates (Greiner . N ü r t i n g e n ; N o . M 2 2 0 29 A R T ) . E a c h serum d i l u t i o n was tested f o u r f o l d . Cont ro l s were r u n for the ca l cu la t ion o f the T G I !)-,„ m l w i t h o u t add i t i ons , o the r con t ro l s con ta ined guinea p i g complemen t and v i r u s w i t h o u t an t ibodies . H u m a n a n t i v a c c i n i a i m m u n e se rum (Vaceinla-b i i l i n ® / I i n m u n o ) served as s t andard in each test . N e u t r a l i z a t i o n u n i t s N U 50 were coun ted accord ing to the m e t h o d o f R E E D and M i : ION en (18) af ter 48 hours .

Results

0 p t i in <* 1 i n c n b a t i o n t i m e an d d o s i s - o p t i m u m of a n t i g e n

The incubation time of culture required to give an optimal transformation-ratio at a given time (day 20-25) after vaccination was assayed in primary vaccinated volunteers (N — 5). Augmented 3 H -Thymidin incorporation could already be observed after 72 hours and reached a maximum after 120 hours of incubation, later on the incorporation ratio declined rapidly. Infectious partially purified vaccinia virus was used as test antigen (Fig. 1). For every preparation of viral

454 • U . K O S Z I N O W S K I , B . V O L K M A N N and R . T H O M S S E N

o

< er * z * ° " g

< z er o u. Ü> z < er h- 1 5 . .

10--

5--

i 1 1 1 J 1 5 72 96 120 1 U 168 foours

INCUBATION TIME

F i g . 1. T r a n s f o r m a t i o n ra t ios a t d i f fe rent t imes af ter a d d i n g 0.2 m l p a r t i a l l y p u r i f i e d infec t ious v a c c i n i a v i r u s an t igen . Mean values o f ( N = 5) l y m p h o c y t e donors 20-25 days a f te r p r i m a r y v a c c i n a t i o n .

antigen the dosis-optimum had to be determined. Figure 2 shows the dependence of transformation-ratio on antigen-dilution using a non-purified infectious cell homogenate (VZO). Whilst concentrated material (protein content 1.5 mg/ml) showed no activity, transformation-ratios increased with dilution of the antigen up to an optimum at a dilution of 1 : 10. Partially purified vaccinia antigen on the other hand induced specific transformations in the undiluted state.

Spec i f i c i ty of v i rus induced l ymphocy te t r a n s f o r m a t i o n

To confirm the virus-specificity of the transformation, transformation-ratios induced by vaccinia antigen (both infectious and UV-in-activated) and 2 control antigens, vesicular stomatitis virus (VSV) and a Vero-cell homogenate were compared. VSV-antigen was used in a concentration that wras effective in inducing transformation of lym-

Cel l -Media ted I m m u n i t y to V a c c i n i a V i r u s • 455

1

I 1 1 . 1 1 1 1 » — >

1 1 1 2 1:5 1:10 1:50 1:100 1:500 1:1000

ANTIGEN DILUTION

F i g . 2. T r a n s f o r m a t i o n r a t i o s o f crude vacc in ia v i r u s in fec ted tissue culture; m a t e r i a l (Vero-cel ls , p r e p a r a t i o n V Z 9 ) . T o x i c effects o f concen t ra t ed m a t e r i a l can be r e m o v e d by d i l u t i o n i n tissue c u l t u r e m e d i u m . L y m p h o c y t e donors ( X — 5), 20-25 days after p r i m a r y v a c c i n a t i o n .

T a b . 1. Speci f ic i ty o f vacc in i a v i r u s induced l y m p h o c y t e t r a n s f o r m a t i o n

Test c o n t r o l V a c c i n i a Vacc in i a Ves icu la r Vero-ce 11 an t igen N o . w i t h o u t ( infect ious) ( U V S t o m a t i t i s c o n t r o l

an t igen i n a c t i v a t e d ) V i r u s ( infect ious)

25 171 1201* n . t . 35 168 27 151 778** n . t , 192 85 40 188 1470* 4 7 4 *̂ 191 n . t . 41 150 1364* 795* 164 145

mean o f 5 values, d . p . m . * < P 0.01 c o m p a r e d w i t h an t igen free con t ro l and Vero-ce 11 an t i gen c o n t r o l

** < P 0.05 c o m p a r e d w i t h an t igen free c o n t r o l a n d Vero-ce l l an t igen c o n t r o l n . t . : n o t tes ted

456 • U . K O S Z I N O W S K I , B . V O L K M A N N a n d R . T H O M S S E X

phocytes from VSV-sensitized guinea pigs in-vitro as shown by earlier experiments (13). The Vero-cell control, equal in protein content to the vaccinia-virus preparation, was run, because the test antigens had been propagated on Vero-cells. As shown in Table 1, only vaccinia antigens were able to transform lymphocytes of vaccinia-sensitized persons.

Trans fo rmat ion induced by di f fe rent vaccin ia ant igen preparat ions

Different vaccinia virus preparations harvested from BHK-21 and Vero-cells were tested as in-vitro antigens. Unrelated to protein content or TCID 5 0/ml. the harvests differed widely in their transforming activity, whereas inactivation was always accompanied with a certain loss of in-vitro transforming activity (Fig. 3).

8

6

different purified form. UV Antigen virus harvests inactivated Behring

F i g . 3. T r a n s f o r m a t i o n a c t i v i t i e s o f d i f fe rent vacc in i a v i r u s a n t i g e n p repa ra t ions , p ropaga t ed on B H K - c e l l s . L y m p h o c y t e donors ( N = 8) . 20-25 days af te r p r i m a r y v a c c i n a t i o n . A n t i g e n 0.2 m l , 1 : 10 d i l u t e d i n m e d i u m , i n c u b a t i o n t i m e 120 hours .

Cel l -Media ted I m m u n i t y t o V a c c i n i a V i r u s • 457

Virus specific i n - v i t r o t r ans fo rma t ion after p r i m a r y vacc ina t ion

17 healthy volunteers were vaccinated with vaccinia virus strain Elstree with simultaneous injection of 1000 I U anti-vaccinia globulin. On the day of vaccination and 4 times later at intervals of 4-5 days blood was taken for assay of lymphocyte transformation with vaccinia antigen (antigen VZ9 propagated on Vero-cells was used throughout this study). The formation of antibodies was tested in parallel. Each test person developed significant vaccinia virus specific transformation. The first specific transformations were seen in the second week after sensitization (Fig. 4). The onset of in-vitro transformation and specific cutaneous reactions did not correlate, the latter were always 3-5 days positive before the beginning of specific lymphocyte transformation. At the time of maximal transformation of lymphocytes no antibodies to vaccinia could be detected by neutralization test.

• — • 1 1 1 • 0 5 10 15

DAYS AFTER VACCINATION

F i g . 4. D e v e l o p m e n t o f cell m e d i a t e d i m m u n e response ( s t r a igh t l ine) a n d o f n e u t r a l i z i n g a n t i b o d y f o r m a t i o n (dashed l ine) af ter p r i m a r y v a c c i n a t i o n ( N = 17). A n t i g e n V Z 9 , i n c u b a t i o n t i m e 120 hours .

Immune reactions after r evacc ina t ion

11 healthy volunteers who had already been vaccinated within the last 1-7 years were revaccinated. Al l persons had established cell-mediated immunity to vaccinia virus and no booster effect on in-vitro reactivity after revaccination was observed (Fig. 5). Simultaneously

4 5 8 • U . K O S Z I N O W S K I , B . V O L K M A N N and R . T H O M S S E N

-500

z

-100

I 1 1 1 1 '

0 5 10 15 DAYS AFTER REVACCINATION

F i g . 5 . T r a n s f o r m a t i o n r a t ios i n l y m p h o c y t e cu l tu res after r evace ina t ion . a) • L y m p h o c y t e donors w i t h specific cutaneous react ions ( N = 1 1 ) . b) O L y m p h o c y t e donors w i t h o u t specific cutaneous react ions ( N - 5 ) .

L i n e w i t h dashes: increase o f n e u t r a l i z i n g an t ibod ies persons successfully re-vacc ina ted (a) .

the sera of those persons who had shown positive cutaneous reactions were tested for neutralizing antibodies. Although there was no augmentation of CM I in-vitro, an increase of the antibody level to vaccinia could be observed.

Discussion

In persons sensitized to vaccinia virus, cell mediated immunity can be demonstrated by lymphocyte transformation test with a good reproducibility. The results are virusspecifie and unspecific cross reactions with an unrelated virus were not seen. Lymphocytes from persons without prior contact to vaccinia virus in no case developed positive in-vitro reactions. The incubation time to obtain optimal transformation with vaccinia antigen corresponds well with the incubation time using other bacterial, cellular and viral antigens for demonstration of CMI in-vitro. Difficulties may appear in the choice of antigen for lymphocyte transformation. In our system commercial vaccines did not work well. Moreover, by using the same vaccine as antigen for in-vitro testing, as used for vaccination, there may be some interference of virus specific immune reactions with cell mediated immune response directed against antigens on the host cell used for virus

Cel l -Media ted I m m u n i t y to Vacc in i a V i r u s • 459

propagation. In experiments with laboratory animals, vaccinia virus worked as an immunological adjuvant to host cell transplantation antigens (1). Therefore, as commercial vaccines were propagated in calves the test antigens were prepared from virus grown in tissue culture of different origin. The harvests differed significantly in their transforming activity not dependent on virus and protein content. Purification of virus particles did not always lead to higher blast formation. Not only viral particles alone, but virus induced cell surface antigens may also be capable of inducing cell mediated immune reactions in-vitro (22).

Since some viruses can transform and replicate in lymphoid cells or depress lymphoid immune reactions (4) the use of inactivated vaccines for in-vitro testing might be advantageous. In our experiments, working with optimal antigen concentrations, we never saw an unspecific inhibitory effect of infective vaccinia virus. There is little known about the onset of delayed type immune reactions after primary vaccination. While several authors could show lymphocyte transformation to vaccinia in immune human subjects (6, 10, 12, 24) there exists only one communication concerning the time course of development. By microscopic counting of blast cells augmented lymphocyte transformation after antigenic challenge in man was observed 20 days after vaccination (20). In vaccinia injected animals enhanced transformation reactions to vaccinia were already seen after 5 days, reaching a maximum in the second week after sensitization (19). In experiments with guinea pigs (13) and rabbits (unpublished data) we also found increased transformation ratios at earlier stages. Delayed type vaccinia-specific cutaneous reactions always preceded the onset of positive in-vitro transformation reactions. This difference may represent the time needed for sensitized cells to reach the circulation: Depending on dose and way of antigen application active lymphocytes can be found first in regional lymphnodes, then in the spleen and several days later in the peripheral blood (23). In experiments with rabbits (19) optimal transformation reactions could be demonstrated only within a relatively short time range after immunization. The time of harvest of lymphocytes for in-vitro investigations may therefore be very important. The early peak of lymphocyte response can also be observed in their effector functions, by testing cytotoxic activities on virus infected target cells (9). I t remains to be determined, what kind of cells are responsible for the immune response later after infection.

In contrast to the well known second set reaction in transplantation experiments we found, apart from an increase of neutralizing antibodies, no significant differences in the proliferation of lymphoid cells of vaccinia immune donors after re vaccination. We suggest, that in immune persons, who show specific blast transformation, the effects of re vaccination on delayed hypersensitivity may be restricted to the

400 • U . K O S Z I N O W S K I , B . V O L K M A N N a n d R . T H O M S S E N

draining lymph nodes and that the generalized effects are small and, unlike the first vaccination, escape the lymphocyte transformation test. Similar results have been obtained by other authors looking for interferon production after revaccination (6). While there was an increase of interferon production in-vitro following addition of vaccinia antigen, no significant alteration of lymphocyte transformation after revaccination could be observed.

Hence it may be of doubtful value to use the lymphocyte transformation test to quantify the vaccinia virus immune status for clinical purposes. However, it may be a useful test in looking for CMI in diseases with impaired delayed hypersensitivity or in cases of uncertain cutaneous and serological reactions after primary vaccination.

Acknowledgement W e t h a n k M r s . G . K Ö H L E R for s k i l l f u l t echn ica l he lp .

Zusammenfassung

I n - v i t r o -Nachweis z e l l u l ä r e r I m m u n i t ä t gegen V a c c i n a v i r u s b e i m Menschen

Z e l l v e r m i t t e l t e I m m u n i t ä t gegen V a c c i n i a v i r u s w u r d e i n v i t r o d u r c h d ie L y m p h o z y t e n t r a n s f o r m a t i o n menschl icher L y m p h o z y t e n un t e r such t . V a c c i n i a -A n t i g e n , v e r m e h r t a u f B H K - 2 1 u n d Vero -Ze l l en , k o n n t e m i t E r f o l g i m te i lweise gere in ig ten w i e auch i m unge re in ig t en Z u s t a n d fü r die I n - v i t r o -Testu n g ver wendet we rden . V a c c i n i a - A n t i g e n - P r ä p a r a t i o n e n w a r e n sowohl i m i n f e k t i ö s e n als auch i m i n a k t i v i e r t e n Z u s t a n d w i r k s a m , die I n a k t i v i e r u n g w a r g e w ö h n l i c h v o n einer gewissen E i n b u ß e an s t imu l i e rende r A k t i v i t ä t begle i te t . Die E n t w i c k l u n g der zell v e r m i t t e l t en I m m u n a n t w o r t nach E r s t i m p f u n g w u r d e bei 17 E r wachsenen un te r such t . Vacciniavirus-spezif ische L y m p h o z y t e n t r a n s f o r m a t i o n e n k o n n t e n i n der zwe i t en W o c h e nach der I m p f u n g i n al len F ä l l e n beobachte t werden . E l f ge impf te Personen w u r d e n h i n s i c h t l i c h ih re r I m m u n a n t w o r t nach W i e d e r i m p f u n g un te r such t . W ä h r e n d die neu t ra l i s i e renden A n t i k ö r p e r anst iegen, w a r eine Z u n a h m e der L y i n p h o z y t e n t r a n s f o r m a t i o n s r a t e n i c h t nachweisbar.

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z y t e n sensibi l is ier ter u n d n i c h t sensibi l is ier ter Personen d u r c h Vacc in i a -A n t i g e n . In fec t ion 1, 29 (1973).

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