DE2119688C3 - Manufacture of live virus vaccines - Google Patents

Description

The subject matter of the invention is a method of manufacture of vaccines with live mumps, flu-, flu-B, cowpox viruses, viruses of the Neweastle disease, Viruses of the Japanese kncephalilis, dengue fever viruses, Rabies viruses, Marek's disease viruses. Rubella, distemper or measles viruses according to the Web culture process. characterized in that / in order to breed the viruses or hatched quail eggs Wachielcnibryofibroblasts are used.

From British Medical Sulclin 25. No. 2. pp. 119-208 (l% 9). are numerous processes / for the production of l.ebendvir>: simpisloffen using chicken and Enicnenibryokulluren known.

The commonly used tissue culture method poses the difficult problem. that depending on üevii. what a supplier for the fabric is used, which the host / hurry accompanying unfavorable I aklorcn often contaminate the culture / hurry, which leads to the inclusion of such unfavorable factors in all finished vaccines. Usually is It is very difficult not in advance not to look into external symptoms ma lifeslicremle infections ties that To discover tissue-producing animals with such undesirable l'akloren. For example, one poses often found in apparently healthy monkeys great ape virus, which / u difficulties in production from l'olioimpfsloff Führl. It is also reported that dal! Discover Newcastic Vircn in tissue cultures of apparently normal Enlenembryofibroblasteii! became. It is also known that the use of Chicken kidneys as a host animal for the breeding of viruses the Marekschen llüliner leaning inevitably with the Danger. that in the vaccines produced against the Mareksche Krankheil llühncrlciikiiniicviren be introduced.

From the above-mentioned round rules, when using the tissue culture method for the preparation of vaccines, care should be taken to eliminate the risk of accompanying unfavorable lactors being introduced Helpful materials in bcschriinklen quantities available require SIML and a complicated handling and umfangrciclK ¹ and complicated analysis.

Task (Scr invention is therefore a method / ur Preparation of vaccines in certain living Viruses that immunize animals and humans are extremely effective and e \ irem iivirulent against viral diseases, thus making them safer vaccinated animals and humans! will.

and into which unfavorable or undesirable | - "actuators are hardly introduced, / u simply manage to do so manageable and easy to implement, and so an economical mass production of virus vac / ine > permitted.

This object is achieved with the invention.

With extensive and intensive research to overcome the disadvantages of the conventional Procedures employing the usual ones in tissue culture are unavoidable, surprisingly found that hatched, i.e., containing embryos, quail cows and tissue cultures from Quail embryo fibroblasts for the cultivation of viruses / ur production of virus vaccines extraordinarily gul ι> are suitable.

By using hatched quail cier or quail embryo fibroblasts, maii can do more than just that Get high quality virus vacancies, but also those that are more suitable due to the low availability '"Materials and the complexity of the method of operation that have arisen so far in an advantageous manner clear out.

The use of incubated waehl eggs (AWE) or tissue cultures from Waehlelembryofibro-. '" > blasten (WEF) bielel the following Korlschritl:

1) A significant number of virus species have an affinity for hatched quail eggs and Gcwebckuhiiren from WEI- "and can be on or in the

, "Named host / hurry in sufficient measure

grow and multiply to produce a vaccine to enable.

2) Both Verwemluni! of hatched quail eggs and glazes from WII- "isl im

;, Contrast to the use of hatched

Chicken eggs are hardly at risk of being infected with vaccines or virus kiiliureii I lühnerlcukemia viruses to bring in.

i) Incubated quail eggs and neck eggs

i "WEI-" are readily available and at low cost

available.

4) With regard to Gcwebektilluren is the number of cells to be obtained from an embryo from a hatched quail egg are about the same

'' like the number of those from a hatched I lühnerci

cells to be harvested. In addition, a greater number of Quails are considered to be bred or kept by chickens, as quails are smaller than chickens. the Zuchtkonirolle with regard to the specific l'alhogenfreilieil (Sl '! -') is easy to stir.

)) Especially in the preparation of vaccines Superior vaccines with a high safety factor can be used against sick people Use of quail produce, which is practically free from chicki-iliological microorganisms contain.

• i · / ur! IrfimliiiifshohL 'becomes niisg \' ftiliii:

On the one hand, it was surprising that the degree of discovery did not use guardians included.

On the other hand, it was even more surprising that

'·· between the mentioned quail and the According to the invention, aiienuicrtcn and multiplied viruses Look at a specific relationship! the one favorable Ailaplion. Attenuation and reproduction of these viruses

admits- Diis is completely surprising, because by no means all Viruses are on every Λπ by Ciewebe or; uif every Λπ win poultry tissue / u multiply. So is /. H. the Maisuera-Siamm, a Röiel tribe. not on the Propagate the chorio-allantoic membrane of the chicken uder to old while he was on the chicken amninion is attenuated to a small extent. An influenza strain multiplies on the C'horio Alleniois membrane, while he is not aware of it.

So there are specific relationships between the virus strain and the host tissue, which are by no means mutually exclusive let predict.

Hence, it means a lucky handle, one Gewebearl to have found both the attenuation and the multiplication of certain Viruses allowed with good results.

The isolation of disease viruses from the materials infected with them and the adaptation, the wax egg: and the cravings; of the isolated viruses are

Table I.

chicken

Λη / iihl der FeMhühner

Methods of the invention, of course, by conventional methods well known to those skilled in the art carried out.

The invention is explained below on the basis of experiments and examples.

Attempt 1

Hiilvers and quails raised by the conventional method are carefully examined for infection with chicken leukemia viruses (hereinafter referred to as "RIF"; see M. Rolle and A. Mayer, Mikrobiologie und Scuchenlehre, I4bb, p. 7J5) Attention is to be paid, and / was particularly, when vaccines with live viruses are to be manufactured. The investigation is carried out according to the method of Rubin (Proc. Nat. Acad. Sci. 4b. 1105, 1960) and the method of Sarma and coworkers (Virology. 23, 51 J, 1964). The results obtained are shown in Table I / w .

Number of animal chickens with a positive Itelund

quail

number of
Tesl quail

RIF antibodies at 5.1

8, -67

RIF in the embryo
RII- in the Ellcmvogel

number of
Quail mil
positive
Finding

163
85

As is clear from Table I / »isl seen. it turns out that most of the adult males contained KIF antibodies, which shows. Most of the chickens were infected with RII- ". In contrast, it could be demonstrated that none of the three adult quails had RII antibodies. The examination showed eagle owls and the internal organs of embryos and adult birds for RII- ' none of the examined adult quails KII- "was detectable. From these results it can be concluded with a fair degree of certainty that the probability of a natural infection of quails with KIFatißor is extremely low.

Attempt 2

One hatched chicken egg and one hatched quail egg each were made using the conventional method subject to dispersion animal contained therein / rush. Then the number of / rush per embryo certainly. The results of these tests are listed in Table II.

Table Il number of / el len K) Emhryoari KO XK) ^- S, ') X K) chicken 7.S x K) ' 8.8 X quail

As can be seen from Table II, I are liihner and Quail bc / üjilich the number of (lurch tissue culture of embryos obtained / rush to roughly equate each other.

Attempt i

Isolation of viruses animal infectious lü'ihnerliihmting oiler Marek's disease (hereinafter referred to as Marek virus oiler MD for short).

The material used to isolate Marek's viruses is lilut or tumor tissue from paralyzes infected chickens. The tumor tissue is treated with trypsin according to the usual procedure, whereupon the parts of the tissue are dispersed. The / rush are tlar.n in culture broth according to Eagle suspended.

For comparison, a 7 to 15 day old quail embryo is processed into an /elldi.ipersion according to the usual procedure. The fibroblasts obtained in this way are ^{plated in a titer of 2.4 10 hours} per Petri dish with a diameter of b cm. The next day the fibroblasts on the trischalcn are each inoculated with 1 ml of the above-mentioned Ulm or tuniorgewcbesuspension, whereupon the preparations are placed in an Urul apparatus. which contains 5% carbon dioxide gas. Cultivated for 4 to 7 days at J7 "C. The results are summarized in Table III, in which" + "a positive and" - "a negative result of the test for Marek viruses isolated in the respective fibroblast cells or for the respective Fibroblast culture-adapted M a ick viruses mean.

/ um comparison are also with chicken kidneys. l. duck embryos and turkey embryos as the culture host are listed.

Table III Used knee No. of the test material KKK ¹ I. vngels cpi: ⁴ i ii ^:> ) Letter + + Chick 13 spleen + + kidney + - + blood + + liver + + Chick u spleen + + kidney -f. ^ - blood + + liver + + Chick 23 spleen -i + kidney + + blood - liver + + Chick 76 spleen + + kidney + + blood + + liver J_ _ | " Chick 105 spleen + + kidney + + blood blood + + Turkey 5 blood _{+ +} Turkey 8

quail
WKK ¹ I.

CPK

Chicken
HNK ¹ )

CPK Il

Annotation:

¹ ) Embryo fibroblast culture.

^:) Wax cmbryofibroblast culture.

¹ I Chicken nicrene culture.

^4> . Cytopalhic Clicks.

■ "") Imniunofluorcs / rcnztechnik.

As can be seen from Table III. Marek viruses were found in decnembryofibroblast culture L ^: L ^: K. Waehtclembryofibrnblastkultiir WEK as well as in chicken nucleus culture HNK. It should be noted that Marck's virus could be isolated by wax embryo fibroblast culture. Another notable valley thing is. that it was found that Marck viruses from the UIu '. infested chickens in a high ratio of 4/5 and from liver. The kidney and spleen were each separated in a ratio of 5/5.

The viruses isolated in this way each have an affinity for cells from duck, quail and chicken embryos. It was also observed that these viruses after cultivation in successive cultures (KultuiTcihcnzüchtung) amenable to further growth was.

As can be seen from Table III / u. was the fact whether the embryo fibroblasts used with r.iaiek-Vircn or Hcipesv; ren infected by the turkey were determined by means of the cytopathic effect and the immunofluorescence / neuroscience. The two Experiments obtained results were further checked ir. With the help of an electron microscope.

H e i s ρ i e I I

Prepared in the manner described in Experiment 3 Quail-. Duck and chicken embryo fibroblasts each i ". Kiiluirbrühc after Eagle in one Concentration of about 5x10 "cells per ml suspended, whereupon 100 ml of each of these cultures in Kiiltur flasks were poured into Roux. Then lets you put them in an incubator one at a time 57 C, whereupon they each with MD viruses from the strain BIKEN C (name of the MI) virus strain, which is im "Research Institute for Microbial Diseases. Osaka Universii ". was isolated and obtained from there can be vaccinated. This was per flask

cmc lmplkullurmcn ^ c mim I ml \ crucndct. Linen Lil ' Milk of the HciiiipliiMj. ' uMiilc ilic KuIlin I "" Minii removed. uoiMiil die JIi der Kolhenw .nid liiiilcndcn / rush mil I hulks' I.osun ^ L'cuiischeii «iirdcii. llicniiil was again pen in losiiiijj after I.ml'Ic im die kolben i: ci: ossen and the / üchtuii}. ' hei S7 C 4 to 7 I; ij; ». · lorlficsci / l. Into the die / elherilndeiUMf; so ueil forljicseliriilen was. dall 4 (1 to 4 ")" / "the fjcsamlcn / hurry veriiiulcrl lullen Then uuiile die / iiclitun ^ heeiulel. N.I bccndelei / Helium ^ were the / rush / iir llcrsiclluii}. ' infi / icrici I nibiMihbioblaslkiillui en \ crwcndcl. those in ΚιιΜιιιΊο sinn: me l.jfile in a concentration min ~> ■ clusters per ml suspend! have been to a mli / ierlc kiiluirsuspension / u increase, du- chitin n; i / iisiii / of a jiceignclen Slahilisalor. / Ii. (ihzerin. fjel.ifjcrt u urile.

| edc of the inli / iles obtained in this way l.mbrMiiihrobl.istkulturcn became. · chinti genial! your Norms luml >> l I.S. Standards lor Λρρπη.il oil V; iccine " iiui \ eiscliiedene (ijrcnschiif lcn unlersuclit. around the Safe for vaccines. Diiiin were they / η hiindelsublichcin Miuck Impfslolf mn lehcuclcn viruses konleklionicrl

Ii c ι s ρ ι e I 2

iJuivn UHcMenmrxoimnim.isikiiiiiir ii K isolated M.Hck viruses. /.H. Miirek viruses muh Sl.imni HIkIN ((^) (M; irok Vu ensUinuii. Isoiierl \ <> m »Keseiirch Instiiii Ie oil Microbiiil Discuses. I limcrsiliii Os.ik.i»: there iiiicli ei hiilllicli) become one (vPiissiijicn- . ib-l'iissiiL'cn · Ivw. 4 (i-! 'iissu ^ enkullurreihen / uchliinj.' unleiuin len. where ius finally important elembiAolibrobhistkiilmr \ eru en d. \ l'I. lieispiell) a 12 l'iiss.iL'cnkulliirreihen / 'Uchluiii! linier \ erwen diitiL' an Inlenciiibiuilibrobliisikultur and a 24-l'iissiij.'en. 21 I \ iss.il'cii. 4h I '.iss.il'cm- or 72-l'iis s.iL'cnkiillun eihcn / iichliini. 'with the use of a W iichlclembiAolihrobl.isikullur unlcr "orlcn.

The bumps obtained in this way cultn'cn cry | ew en in eggs in the ncispici ι nese iiiici'eneu wise treated. to get an inh / ierle kiilliii ispension / u obtain. The, ml in this way received illcncn kiiltiirsuspen Sions become iiiieli / iiL'iibe of i / ceit'tielcn Suibilis.i lors. /.H. (iheerin. ^ 'ehigert. wor.iul their I iihiijkcit. Immunize chickens horny M.irckschc kr.inkheil / u, unlearning w ills.

The above described vaccines are given own / en n.icli eiiiwochiL'ei l.iijjeruiiL 'at 4 (and myself iiii. , Hei l.iiL'eriini; at very deep i'ii lempciiitu regret. /.IV iiiich J? Mon, ilii_'cr l., Ii.'en nj: at minus 70 (. cmc iiusreieheiiele lninninisicruni; s.ikti \ iliil.

The 'dragonfly l \ μιΐΐ! before l.iijcnschiiiten the. how \ οι see iiel described, received In ι pistol Ie w leeler.

Dragonfly | \

\ he -lent-

/ aiii elei All animals with verucmk ic ν eggs Mrulentc.il Chick St.mim MMi \ l.irek- \ iren ec inipl'len chicks (lace I

Hnimpl'imi: ik- InipKlolK

Anelort'olpcnelen cultures overlaid
(Kiilturreihen / iKtiluncr I Kiikenl I'.icei

All of the test 1 I.ieillpkcit ikr

kiiken heim I rkr.iiik.inL · .in

Kont.ikl with M.irckVliL'r

hel'jllenen Kiiken Kr.inklie '.')

HXi
B-

- ¹ II

I ¹ '

3 pp

5 (1

50

50

BIKIN (Stamm I IK I 2 -

BIKIN C strain FEK12 -: K 4 (i

WI K 72

BIKIiN Q stem WEK 6

BIKING 0

Strain WEK 36

BIKING 0

tribe

WEK 46

BIKEN C

Strain EEK 12 -WEK 24

4 pp

18th

14/2 (1 (7 (1) (1/2 (1

(l / l ¹ )

7/20 (35 ', I.

2/50 (4%)

0/50

0/50

Experimental /.ilil the

MIlT tier mil I

group

\) Comparison

used the ν irulcnlcn

Kiikcn

-IS

Sliinini by
Marek viruses
iiiipllcii chick
(I.ige)

ιηιηιρίυημ (Ils Implslolls

Breeding in aul'ein- all der

inoculated in other cultures
(Kullurreihenzüchtiing) ') chicks (days)

HIKIN C

tribe

1IK 12 VVI-K 27

IiIKI N C

tribe

ΐ: Κ7_ '· Wl K I

H)

Age of l'estkiiken at
Contact mil
IK ¹ IaIk ¹ IiLMi chick
(location)

4 ')

Frequency of
Disease
Marek's seer
Disease ¹ I.

IAtH (2 ",

0/20

Annotation.
'A: Because. (E Leghorn.
• H: DeKaIh IhI.

'l-.s were each I). 2 ml linplsloll in the Mauchhühl · geimpl'l. The surveillance was in each case only lorlgeliihil. until the test animals were 2Λ weeks .dt.

The virulent strain of the Marek virus is a through / üchtiim: in mutually overlapping cultures after home of Chicken puff caves with mint from trunk raised with chickens included.

) "HIKIiN (strain" and "UIKLN () strain" are the drawings of MD-Virenslänimcn. Which were isolated in the "Research Institute Tor Microhial Diseases. Osaka Illnivcrsiliit" and are available there Hl '^ ml. Given in I'laque-Hildungs-Imhcilen (l'll ^! ). The numbers attached to the abbreviations Ii-K and Wl-K refer to the number of passages during the breeding.

) The lestluihner were held together with several chickens in the same (iellügelh.nis, vim which we know; ir. because they were afflicted with ransom.

') The infected and loose chickens were each excised and examined Diagnosis of the disease made from chicken lair

As can be seen from I ahelle IV / ii, / cigle der durch Kultiirrcihcn / üchtung mn high l'assagen / ahl (/. H. with 4 J-I'assageii) obtained using vim MI-IK Vaccine a superior immunization effect against Mareksehe Krankheil. Also animal by classification with a few l'assagen using LMK and subsequent Kulturrcihcii / üchtting with about 20 or more l'assagen obtained using WHK vaccine had a superior · that of those from by culture series / üchlung with high passages / ahl internal use of WKK obtained alone Vaccine-like immunization activity against Mareksehe disease. It is thus statistically proven that that Marek viruses by culture series / üchlung using MMK and / or WMK im essential avirulenl can be made and that the cultures obtained as a Marek vaccine are usable and valuable.

Example J

7 to 12 day old quail embryos are decapitated, amputated and gutted and then minced. The resulting mass is mixed with 0.2% aqueous trypsin solution and then treated according to the usual method for dispersing the cells. The dispersion obtained in this way is centrifuged at low speed to remove the aqueous phase and then in 5% calf serum. 0.5% hydrolyzed lactalbumin and 50 ug / ml erythromycin-containing Earlc-L solution in a concentration of 1.0 to 1.5 10 ⁶ cells per ml suspended.

From the suspension obtained in this way, each K) OmI is poured into Itmiv flasks with a capacity of K) OOmI. Then liild the piston 2 position in Yl C in a llrulapparai stand, whereupon the above-mentioned Kiilturlösimg removed and the adhering to the flask wall cells are washed free mil I lank solution of its (ichall of calf serum. Then, the I ibro blastkiilluren in are the Kotix flasks each with I ml of a suspension infused with measles virus (measles virus from the strain TANAMKflie / drawing of a measles virus strain which was attenuated by culture series using C'horio-allantoic membranes from available II ¹ H chicken eggs] Then add 100 ml Magle-Kullurbrühc / ii and cultivate the cultures 4 to 4 lay further at ih C. After the end of cultivation, the virus suspensions are taken and cleaned by removing or filtering the culture cells Virus suspension (measured using human amniotic cells is 10 "'to 10"'"TCID - ,,,,,, [TCIDin = Infection of 50% cultured tissue bekultur]) and is sufficient for the production of dry grain vaccine.

The viruses mentioned above are also subjected to 10-pass culture in series. With each passage of the culture series there is a virus suspension with approximately equivalent and constant infectivity.

The adaptability of the virus suspensions obtained in this way is described in Tabciic ili Wise checked. The suspension is then turned into commercially available measles vaccines containing live viruses assembled.

Il

Ii c i s ρ i c I t

• A tissue culture is produced from a Waelilelembrvo in the manner described in example i , the iliinii with the while; above-mentioned measles virus from vaccinated St.innn TANAHI- ti net, mschliellcnd a Kiilliirreihcn / is subjected üchmng. The obtained Virussiispeiisioii in / l'ehiiten assage the Kiilliirreihen / üchtung winl mil J ¹ VIi arginine. Vn sucrose and hydrolysed gelatin; the stabilizers are added. The resulting suspension is poured in a certain volume into small halls and freeze-dried immiltclb.ir .indert iiiul after Irocknen ΙΟ ^'1 f' ⁽¹ IIWmI look at me n;. I I .iigcrung one year at 4 (only Ii ... ...-, .., ι, ι ι.. ;., ι ,. ,, .. "; t uvt ■ ι γ ί 11;.,. ι ι ·»; .. i .'.- ι, .. ι

II.U'l.'Ml.L..l.lll <t. VItIf. Ulli II «IV ll ', ₍ | -llll. I'll. I.lim.

ment of the lnickliosil; il k: imi thus ills quite .iiisge. / cichnel be / eichnel become. This is what you can expect. That is, the linplstoHpraparalc that have been granted a fiduciary protection are very stable and can therefore be stored in a hurry.

Comparative experiment 4

Measles virus from strain ΤΛΝΛΙ1Ι (see example i). that by Ktilliirreiii / üchlung under Veruendiing \ om (borio / \ lhintois Memlirwiien from iingebnilelen lliih-

l \ ibelle V

nereicrn st; irk; ibgescl "wächl became / ur Mersiellung von / wci different types of Virus suspensions. mis those / white types of implants liergeslelll, an I -l \ iss; igcn / iichlung b / u. A cooling series / iichtiing mil IO l ': iss; igen subject. These two Iniplsloffsorlen become unleieii children as well as with a vaccine (C AM vaccine slolf), the mis viruses liergeslelll was that of C'horio-Alhm loisMembiiinen of; in-fried chicken eggs. which with the above mentioned Mnsernviren vom Sliimm IANAHI. had been vaccinated, winches won.

The CAM vaccine obtained in this way is examined with regard to its variability with the vaccine in the manner described in Example I. With the i | u.ilili / iencn vaccine win! em 8 months to i | ahre everything Child not Masernanliköiper vaccinated. The insurance results are given in Table V / iisamnienge

As can be seen clearly from labelle V / ii. Distinguish the two using a tissue culture from quail telemlyofibrobias vaccines obtained from the CAM implant at the time of clinical response. eggs on () riginalslamm eggs above two \ VLT: n of vaccines and derives from Chorio Allaniois membranes of angebriile th quail eggs, not in a significant way, and have a sufficient lnimunisierungsakli \ iliii on.

lerkunlt dos Inipl'sliilVcs

Inlckliosiliil
ΙηιρΓηιοημο
(Ιορι, ιΓ (ΊΙ) ,,, Ι

/; ilil (lor μο-iin | ilton Kiiulor number of Kiiulor number of children, ie number of

niil / iinahnic

spe / illsclicn

antibody

the

to which a nudge
dor body temperature uul more than
.V) (entry

Children mi!
Aiisschliiu

(iewebekiillur from 3.7

Alternate embryonic fibro-

blast. l-l'assagen-

/ direction

(iewehekullur from 3.5

VVaehlelembryollbro-

bliisten. lO-l'nssngenkullur-

reiheti / üehliinii

Koillrollversueb .1,4

(CAM implsloir)

12 12

H e i s ρ i e I)

Incubated quail eggs are infected with measles viruses b / w. Infltien / a-A.iViruses inoculated. The received Vac / ine are incubated by inoculating

Table Vl

Chicken eggs with measles virus b / w. Influenza virus vaccines compared. The results of this Experiments are shown in Table VI.

Virus type
Measles')

Cultivation time (days)
2 3

Influenza Ar)

Infbkliosilät 5.0 5.3 5.6 6.0 8.5 Quail 4.2 4.7 5.5 5.5 6.5 8.5 Chicken 4.0 Annotation:
') log |., TCH:>, _n / ml
') logmEIDsii / ml

14th

The data listed in the table above u order, determined on the basis of the following experiments:

For the production of mask vaccine, the Iruchtsaek of incubated kings is inoculated with all Kmbryos with 0.1 ml of the suspension of the aforementioned TANAIiI strain for 7 days and then cultivated at 3: 3 ° C. for 7 days. For the production of influenza viruses, the Chono-Allanlois-Menibran (CAM) is inoculated for 8 days in all incubated kittens with influenza viruses of strain A> and then cultivated at i / C for 5 days. In given time intervals !! If viruses are removed and virus suspensions are processed, they are then examined for their infectiousness. The vinisaiis boils obtained from the use of hatched quail eggs are just as good as those obtained from the use of incubated quail eggs.

ι ϊ c i ** ρ ί ο ι ί ι

In de: in examples 3 to "> described manner farlen hatched quail eggs and (iewcbekuliiiien from dialing-cryolibrublastcn with \ ersehiedenen Virus arenas vaccinated. The growth and the multiplication the viruses are determined by known methods.

The results of these experiments are given in the table VII listed.

As can be seen from this. are branded Quail eggs and weave cultures from quail biofibroblasts are superior in terms of Atlaplion and multiplication of various viiusarls. In addition, as already mentioned, comes with

Table YII

hatched quail eggs untl (iewebekulluren from quail embryo fibroblasts tlas kiiischleppen resp. Infiltration of unfavorable actors. especially lousy leukemia. and hatched quail eggs are not suitable for use Wax embryo fibroblasts are good for the cultivation of high quality virus vaccines with live ones Viruses.

Ks were the following virussiiimnie in the given doses:

Virus mute

lot

rubella Miilsuii rii 0.5 ml / dose mumps Urabe 0.5 ml / dose Influenza A. Aiehi 0.5 ml / dose Inlluenzii Ii Kagoshima 0.5 ml / dose Amazement by I lüliner-
embryo
palJler trunk 0.5 ml / dose Neweaslle l-'udiii (Iu-I) 0.5 ml / dose Japanese
Ephalitis M1.-I7 0.5 ml / dose Dengue fever Dl N-Ii 0.5 ml / l ¹ OSJs rabies CKA-I 0.5 ml / dose Marek-schc
illness IiIKIN C-. " ¹ 0.5 ml / dose

Riitelil

Incubated (k-uebe-Kullur (iewebe-Kullur Ciew ehe-Kiilliir Mclhoik '

Wachlole eggs from Wachlcl- from chicken- from Allcii-

einhryollbm- embryo urbation

blow hlaslon

3.5 3.5 4.0

(iewcbe cultures from kidneys of green monkeys

mumps

7. (1

7 .:

7.0

(irippe A _: P.5 1.2 / ^ 7.> (! rib H SO 7.5 1.2 (1.0 distemper 4.1 4.0 .i.S. 5.0 Kuhpoeken SO 7.0 6.5 - Newcastle 8.0 SO 8.2 - Japanese
Finccphalilis 7.8 P.5 7.5 P.5 Denguelleber 4.5 5.0 4.5 5.0 rabies 7.3 4.0 .1.8 - Marek-sche
illness 2.2 4.3 3.9 -

Ciewebekulluren from Iliihrvrembryo-

llbroblaslen

(1OgI ₁₁ TCM) ,,, / ml) ''

Hatched chicken egg

(lot;, KID ^ / min

Fish egg

(logi "Kll)," / ml)

(iewehekullur from llühneremhryo-

Hbmblaslen

ΙΙορι ,, ΡΙ-'υ / ιηΙ) " ¹ )

Hatched chicken egg

(log ", POITi / ml) ⁵ )

Hatched chicken egg

Tissue culture from chicken embryo

llbroblasts

(log ^ PITJ / ml) ⁴ )

mouse

mouse
(log _m LI) ₅ "Zml)

Tissue culture from cone embryo broblasts

Claims (1)

9 688 Claim: Process / ur manufacture of vaccines with Live mumps, Crippe-A, Influenza-B, cowpoke viruses, viruses of Newcastle disease, viruses of Japanese encephalitis, dengue virus, rabies virus, Marek's disease viruses, rubella, distemper or measles viruses after the tissue culture process, characterized in that quail eggs or quail embryo fibroblasts are used to breed the viruses.