Abstract
Antigen Used for Immunization
The immunogen (canine distemper virus [CDV] Snyder Hill strain) was stored in our laboratory.
Method of Immunization
For the first immunization, a group of 6-week-old female BALB/c mice (n = 4) were injected with 200 μL CDV Snyder Hill plus an equal volume of complete Freund's adjuvant (Sigma-Aldrich, St Louis, MO) through hypodermic injection at four different sites on the back.(1) Once every 2 weeks, another hypodermic immunization was administrated by using 200 μL CDV Snyder Hill emulsified in a 1:1 ratio in incomplete Freund's adjuvant (Sigma-Aldrich). After three immunizations, an equal amount of antigen without adjuvant was injected into mice through intraperitoneal injection for the boost.
Parental Cell Line Used for Fusion
Mouse myeloma Sp2/0.
Selection and Cloning Procedure
The culture supernatants were tested by indirect fluorescent assay (IFA) with CDV infection Vero cells.
Heavy and Light Chains of Immunoglobulin
Specificity
Five MAbs showed good reaction with CDV, and 1A4 showed specific reaction against CDV-H protein, 1G5 and 6E11 against CDV-F protein, 2H11 against CDV-P protein, and 6D9 specific to CDV-N protein.
Specific Antigen Identified
MAbs 1G5, 6E11, 2H11, and 6D9 were reacted with both wild and vaccine strains, whereas MAb 1A4 only reacted with CDV vaccine but not with wild strains by IFA test. Only MAb 1A4 holds neutralizing activity to vaccine strains of CDV because it could completely neutralize 100TCID50 CDV virus at 1:64 dilution. The other four MAbs have no neutralizing activity to wild and vaccine strains of CDV.