Quantification of rotavirus-like particles by gel permeation chromatography.
There is a lack of accurate and practical methods that require only small amounts of sample for quantifying virus-like particles (VLP). In this work, gel permeation (GP) HPLC was used to quantify double-layered rotavirus-like particles (dlRLP) produced in insect cells. The proposed methodology utilized two columns in series (pore sizes of 200 and 50 nm) and had a high precision (relative standard deviation below 5%). GP-HPLC not only allowed the routine quantification of dlRLP, but also of assembly intermediaries and other viral structures present in the samples. For the first time, kinetics of dlRLP accumulation could be followed. This methodology is valuable for designing new production processes and for optimizing dlRLP monitoring.
Mena
Jimmy A
JA
Departamento de Medicina Molecular y Bioprocesos, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Apdo. Postal. 510-3, Cuernavaca, Morelos CP. 62250, México.
Ramírez
Octavio T
OT
Palomares
Laura A
LA
D000818:Animals; D000956:Antigens, Viral / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism; D002470:Cell Survival; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient / Q000379:methods; D002586:Cesium; D002712:Chlorides; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007313:Insecta / Q000166:cytology / Q000378:metabolism / Q000821:virology; D007700:Kinetics; D008854:Microscopy, Electron; D010316:Particle Size; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015203:Reproducibility of Results; D012401:Rotavirus / Q000235:genetics / Q000302:isolation & purification* / Q000648:ultrastructure; D013696:Temperature
D000956:Antigens, Viral; D036022:Capsid Proteins; D002712:Chlorides; D011993:Recombinant Fusion Proteins; C442545:VP2 protein, Rotavirus; C061332:VP6 protein, Rotavirus; D049452:Green Fluorescent Proteins; D002586:Cesium; C028019:cesium chloride