A comparative analysis of the methods for titering adenoviruses.
- Author:
Peng-yu SUN
1
;
Yan-ling ZHANG
;
Yu-ming JING
;
Xin-ji ZHANG
;
Zhe-huan ZHANG
;
Cheng-yao LI
;
Bai-hong CHEN
;
Wan-long TAN
;
Hong-wei LI
Author Information
- Publication Type:Journal Article
- MeSH: Adenoviridae; isolation & purification; metabolism; physiology; Capsid Proteins; metabolism; DNA, Viral; isolation & purification; Green Fluorescent Proteins; metabolism; Real-Time Polymerase Chain Reaction; methods; Viral Plaque Assay; methods; Virus Replication
- From: Journal of Southern Medical University 2011;31(2):234-238
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo compare different methods commonly used for titering adenovirus and analyze the advantages and limitations of each method.
METHODSFour recombined adenoviruses (Ad-G-AT2R-EGFP, Ad-CMV-EGFP, Ad-mif-shRNA-EGFP and Ad-CBA-GFP) were amplified and purified, and each was titered by optical absorbance, real-time PCR, green fluorescent protein (GFP)-labeled method, immunoassay, and cytopathic effect (CPE). The results were then comparatively analyzed.
RESULTSNo significant difference was found in the titer amounts derived from GFP-labeled method, immunoassay, and cytopathic effect method (P>0.1). A positive correlation was noted in the titer amounts determined by real-time PCR and immunoassay (r=0.965), even though the value (vg/ml) obtained by real-time PCR was 10 times higher than that by immunoassay (ifu/ml).
CONCLUSIONGFP-labeled method and immunoassay allow rapid determination of the adenoviral titer. Real-time PCR can not directly determine the real infectious titer of the adenovirus, but the result is well correlated to that of immunoassay and reflects, though indirectly, the actual infectious titer of adenovirus. Considering the procedural convenience and shorter time consumption, real-time PCR is still a practical method for adenoviral titration.
