Development of Indirect Immunofluorescence Technique for the Identification of MRC5 Working Seed Cell

Document Type : Original Articles

Authors

1 Department of Quality Control, Razi Vaccine & Serum Research Institute, Agricultural Research, Education and Extension Organization, Karaj, Iran

2 Department of Human Viral Vaccines, Razi Vaccine & Serum Research Institute, Agricultural Research, Education and Extension Organization, Karaj, Iran

3 Department of Quality control, Razi Vaccine & Serum Research Institute, Agricultural Research, Education and Extension Organization, Karaj, Iran

4 Department of Physical Education and sport, Faculty of Physical Education and sport, Kharazmi University, Karaj, Iran

5 Department of Health Management, Razi Vaccine & Serum Research Institute, Agricultural Research, Education and Extension Organization, Karaj, Iran

Abstract

Diploid and continuous cell lines are used to propagate viral vaccines. At Human Viral Vaccine Department of Razi Vaccine and Serum Research Institute, MRC5 diploid cell is used for the development of live attenuated measles, mumps, rubella, and three types of poliovirus vaccines.  Additionally, three continuous cell lines (i.e., RK13, HeLa, and Vero) are applied in quality control tests. Accordingly, cell cross-contamination can occur at cell culture labs, hence controlling the identity and specificity of cells is essential. Indirect immunofluorescence is a sensitive, specific, and simple test for cell identification. The present study was designed to develop the in-house indirect immunofluorescence test (IIF) as follows: homemade polyclonal anti-MRC5 serum was prepared in rabbits, and cross-reactive antibodies to RK13, HeLa, and Vero cells were eliminated. The diploid and continuous cell lines were fixed on Teflon slide using cold methanol and acetone. The reproducibility of the in-house IIF test was evaluated using the agreement Kappa test.  The purity of the three batches of MRC5 working seed cell at Human Viral Vaccine Department of Razi institute was verified using IIF and no contamination with continuous cell lines was detected.

Keywords

Main Subjects

Article Title [French]

Développement d’une technique d'immunofluorescence indirecte pour l'identification de la cellule de semence de MRC5

Abstract [French]

Des lignées cellulaires diploïdes et continues sont utilisées dans la productiondes vaccins viraux. Au département des vaccins viraux humains du Razi Vaccine et du Serum Research Institute, la cellule diploïde MRC5 est utilisée pour la propagation de la rougeole atténuée vivante, des oreillons, de la rubéole et de trois types de vaccins antipoliomyélitiques. De plus, trois lignées cellulaires continues (RK13, Hela, vero) sont utilisées dans les tests de contrôle qualité. Parconséquent, la contamination croisée de cellule à cellule peut se produire dans les laboratoires de culture cellulaire et le contrôle de l'identité et la spécificité des cellules est essentiel. L'immunofluorescence indirecte représenteun test sensible, spécifique et simple pour l'identification des cellules. L’objectif de cetteétude était de développer untest d'immunofluorescence indirecte (IIF) interne. A cet effet, un sérum anti-MRC5 polyclonal a été produitchez le lapin et les anticorps réactifs aux cellules RK13, Hela et Vero ont été éliminés. Les lignées cellulaires diploïdes et continues ont été fixées sur une lame de Téflon en utilisant du méthanol froid et de l'acétone. La reproductibilité du test IIF interne a été évaluée en utilisant le tst de concordance Kappa. La pureté des trois lots de cellules semencières MRC5, utilisés par ledépartement de vaccin viral humain de l'institut Razi, a été vérifiée en utilisant un test d'immunofluorescence indirecte et aucune contamination avec des lignées cellulaires continues n'a été détectée.

Keywords [French]

  • MRC-5
  • Cross-contamination
  • IIF
  • Test de Contrôle de Qualité

References

Barallon, R., Bauer, S.R., Butler, J., Capes-Davis, A., Dirks, W.G., Elmore, E., et al., 2010. Recommendation of short tandem repeat profiling for authenticating human cell lines, stem cells, and tissues. In Vitro Cell Dev Biol Anim 46, 727-732.
Bloemberg, D., Quadrilatero, J., 2012. Rapid determination of myosin heavy chain expression in rat, mouse, and human skeletal muscle using multicolor immunofluorescence analysis. PLoS One 7, e35273.
Brouns, I., Van Nassauw, L., Van Genechten, J., Majewski, M., Scheuermann, D.W., Timmermans, J.P., et al., 2002. Triple immunofluorescence staining with antibodies raised in the same species to study the complex innervation pattern of intrapulmonary chemoreceptors. J Histochem Cytochem 50, 575-582.
Buchwalow, I.B., Minin, E.A., Boecker, W., 2005. A multicolor fluorescence immunostaining technique for simultaneous antigen targeting. Acta Histochem 107, 143-148.
Buehring, G.C., Eby, E.A., Eby, M.J., 2004. Cell line cross-contamination: how aware are Mammalian cell culturists of the problem and how to monitor it? In Vitro Cell Dev Biol 40, 211-215.
Capes-Davis, A., Theodosopoulos, G., Atkin, I., Drexler, H.G., Kohara, A., MacLeod, R.A., et al., 2010. Check your cultures! A list of cross-contaminated or misidentified cell lines. Int J Cancer 127, 1-8.
de Oliveira, A.P., de Castro, M.C., de Almeida, A.F., Souza Mde, A., de Oliveira, B.C., Reis, L.C., et al., 2013. Comparison of flow cytometry and indirect immunofluorescence assay in the diagnosis and cure criterion after therapy of American tegumentary leishmaniasis by anti-live Leishmania (Viannia) braziliensis immunoglobulin G. J Immunol Methods 387, 245-253.
Freshney, R.I., 1993. Culture of Animal Cells, Wiley.
Frisch, J., Houchins, J.P., Grahek, M., Schoephoerster, J., Hagen, J., Sweet, J., et al., 2011. Novel multicolor immunofluorescence technique using primary antibodies raised in the same host species. Methods Mol Biol 717, 233-244.
Hanly, W.C., Artwohl, J.E., Bennett, B.T., 1995. Review of Polyclonal Antibody Production Procedures in Mammals and Poultry. ILAR J 37, 93-118.
 Jacobs, J.P., Jones, C.M., Baille, J.P., 1970. Characteristics of a human diploid cell designated MRC-5. Nature 227, 168-170.
Johnen, G., Rozynek, P., von der Gathen, Y., Bryk, O., Zdrenka, R., Johannes, C., et al., 2013. Cross-Contamination of a UROtsa Stock with T24 Cells – Molecular Comparison of Different Cell Lines and Stocks. PLOS ONE 8, e64139.
Koosha, S., Fesharaki, M., Rokni, M.B., 2004. Comparison of enzyme-linked immunosorbent assay and indirect immunofluorescence assay in the diagnosis of human strongyloidiasis. Indian J Gastroenterol 23, 214-216.
Masters, J., 2002. False cell lines. Int J Cancer 99, 154.
Reina, J., Ballesteros, F., Ruiz de Gopegui, E., Munar, M., Mari, M., 2003. Comparison between indirect immunofluorescence assay and shell vial culture for detection of mumps virus from clinical samples. J Clin Microbiol 41, 5186-5187.
Stills, H.F., Jr., 2005. Adjuvants and antibody production: dispelling the myths associated with Freund's complete and other adjuvants. ILAR J 46, 280-293.
Suzuki, T., Tate, G., Ikeda, K., Mitsuya, T., 2005. A novel multicolor immunofluorescence method using heat treatment. Acta Med Okayama 59, 145-151. Neospora caninum by different diagnostic techniques in Mashhad, Iran. Parasitol Res 100, 1257-1260.
Zachary, J.F., McGavin, M.D., 2013. Pathologic Basis of Veterinary Disease-E-Book, Elsevier Health Sciences.
Archives of Razi Institute
Volume 73, Issue 1
March 2018
Pages 39-44

Files

Share

How to cite

Statistics