Monoclonal antibodies for Streptomyces lividans and their use for immunomagnetic capture of spores from soil.
Monoclonal antibodies were produced to Streptomyces lividans spore surface antigens. One particular hybridoma cell line, 43H6, produced a monoclonal antibody that reacted exclusively with Streptomyces cluster group 21 in an enzyme-linked immunosorbent assay (ELISA). Antibody 43H6 was found to be of subclass IgG1, kappa light chain. Western blot (immunoblot) analysis revealed that 43H6 recognized a major outer spore polypeptide of about 37,000 Da. The epitope was stably maintained in S. lividans spores over at least seven sporulation cycles on laboratory medium and for at least 14 weeks in sterile soil systems. The species group specificity of antibody 43H6 was exploited in the development of an immunocapture technique for the isolation of streptomycetes from soil. Magnetic beads coated with antibody 43H6 were mixed with soil samples seeded with S. lividans spores. Spore-bead complexes were recovered using magnets. Treatment of beads with blocking agents and the inclusion of detergents in the recovery system lessened non-specific binding of spores to beads and improved recovery. In buffer solutions decreasing the spore concentration increased the recovery values for a fixed bead concentration. At a spore concentration of 5 x 10(7) ml-1 the recovery was 4.3% whilst at 5 x 10(2) ml-1 it was 76% for a fixed bead concentration of 0.6 mg ml-1. Using a bead concentration of 2 mg per 10 g soil, approximately 30% of the target spore population of 10(6) c.f.u. was recovered from sterile soil and 4% from non-sterile soil. This method offers a rapid means of selectively recovering and concentrating Streptomyces spores from soil samples.
Wipat
A
A
School of Biomolecular Sciences, Liverpool John Moores University, UK.
Wellington
M H
MH
Saunders
V A
VA
D000907:Antibodies, Bacterial / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology; D005780:Gelatin / Q000494:pharmacology; D007074:Immunoglobulin G / Q000276:immunology; D018189:Immunomagnetic Separation; D008894:Milk Proteins / Q000494:pharmacology; D011092:Polyethylene Glycols / Q000494:pharmacology; D012710:Serum Albumin, Bovine / Q000494:pharmacology; D012988:Soil Microbiology; D013171:Spores, Bacterial / Q000302:isolation & purification; D013302:Streptomyces / Q000276:immunology* / Q000302:isolation & purification
D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D000939:Epitopes; D007074:Immunoglobulin G; D008894:Milk Proteins; D012710:Serum Albumin, Bovine; C024062:nonidet; D011092:Polyethylene Glycols; D005780:Gelatin