Establishment of hybridoma cell lines producing monoclonal antibodies against hepatitis B virus surface antigens (a, d, and r) and development of sensitive ELISA diagnostic test.
A new treble-coated enzyme-linked immunoadsorbent assay (ELISA) kit of detecting Hepatitis B virus (HBV) surface antigen subtypes a, d and r (HBsAg-a, -d, -r) was developed by using four established hybridoma cell lines, of which two specifically secrete monoclonal antibodies (MAbs) against HBsAg-a (anti-HBsAg-a), one against -d (anti-HBsAg-d), and one against -r (anti-HBsAg-r). The approach of hybridoma cell lines' establishment were by fusing myeloma cells (SP2/0) with splenocytes from BALB/c mice immunized with a mixture of HBsAg-a, -d, -r. The ascitic MAb productivity of the four cell lines was at the titres of 1:10(6)-1:10(8). A treble-coated ELISA based HBV diagnostic kit was developed for detecting all of the three responding subtypes of HBsAgs. A 96-well ELISA microplate was coated with anti-HBSAg-a, -d, -r at a ratio of 3: 1: 0.5, with a horseradish peroxidase (HRP) conjugated anti-HBsAg-a as the labelled antibody. For clinical application, the new developed diagnostic kit detected HBsAgs of adr, adw, ayr, and ayw at a rate of lower than 0.25, 0.25, 0.5, and 0.5 ng/mL, respectively. Results indicated that this kit was more rapid and sensitive than that other current ELISA-based kits coated with a single MAb (e.g., anti-HBsAg-a).
Tang
X
X
Beijing Biological and Immune Reagents Center, PR China.
Zhang
X
X
Xu
H
H
D000818:Animals; D000911:Antibodies, Monoclonal; D004355:Drug Stability; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006509:Hepatitis B / Q000175:diagnosis*; D006510:Hepatitis B Antibodies; D006514:Hepatitis B Surface Antigens / Q000276:immunology* / Q000302:isolation & purification*; D006801:Humans; D006825:Hybridomas; D051379:Mice; D008807:Mice, Inbred BALB C; D011933:Reagent Kits, Diagnostic; D012680:Sensitivity and Specificity
D000911:Antibodies, Monoclonal; D006510:Hepatitis B Antibodies; D006514:Hepatitis B Surface Antigens; D011933:Reagent Kits, Diagnostic