A simplified multiwell plate assay for the measurement of hepatitis A virus infectivity
| Autor: | David L. Krah |
|---|---|
| Rok vydání: | 1991 |
| Předmět: |
Serial dilution
viruses Bioengineering Biology Applied Microbiology and Biotechnology Virus Cell Line Tissue culture Antigen Virology Potency Humans Hepatitis Antibodies Hepatovirus Antigens Viral Pharmacology Infectivity General Immunology and Microbiology virus diseases Viral Vaccines General Medicine Titer Cell culture Evaluation Studies as Topic Biotechnology |
| Zdroj: | Biologicals : journal of the International Association of Biological Standardization. 19(3) |
| ISSN: | 1045-1056 |
| Popis: | A standardized multiwell plate assay (MWPA) was developed to provide a simple in situ measurement of hepatitis A virus (HAV) infectivity titers. Following attachment (4 h, 35 degrees C) of serial 10-fold dilutions of HAV strain CR326 F (variant F') to confluent MRC-5 monolayers in 24-well plates, cultures were overlaid with maintenance medium and incubated for 35 days at 35 degrees C with weekly medium replacement. Cells were fixed with 90% acetone and HAV antigen was quantitated by reaction with a radio-iodinated anti-HAV serum. Measurement of virus infectivity was based on the amounts of specifically bound and eluted radiolabelled antibody. Virus titers (50% tissue culture infectious doses/ml, TCID50/ml) were calculated using the method of Reed & Muench (Am J Hyg. 1938; 27: 493-497), with wells producing cpm values greater than 2.1-times that of uninoculated controls considered positive. The use of a virus standard provided an estimate of the test variability (+/- 5% in Log10 TCID50 units). The MWPA offers a significant savings in time and reagents, and is proposed as a standard method for the simple and reliable measurement of the potency of HAV vaccine preparations. |
| Databáze: | OpenAIRE |
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