The use of streptavidin-biotin interaction for preparation of reagents for complement-dependent liposome immunoassay of proteins: detection of latrotoxin.
We have developed liposome sensitization by a protein, latrotoxin (LT), using immobilization of biotinylated LT via streptavidin with biotinylated phosphatidylethanolamine contained in liposomes. The use of such liposomes in the complement-dependent homogeneous liposome immune lysis assay (LILA) has allowed us to detect in the test sample as little as 2 micrograms/ml of polyclonal and 50-100 ng/ml of monoclonal IgG and IgM antibodies to LT. LT concentration in solution was determined by inhibition of immune lysis by free LT. The sensitivity of the LT assay varied from 1 x 10(-9) to 5-50 x 10(-9) M when antiserum (polyclonal antibodies) and monoclonal antibodies to LT were correspondingly used. The results show that a streptavidin-biotin spacer can be used to immobilize protein antigens on liposomes for a subsequent application in LILA. The suggested technique greatly simplifies the sensitization procedure and extends the applicability of the LILA.
Pashkov
V N
VN
Branch of the M. M. Shemyakin Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow Region, Russia.
Tsurupa
G P
GP
Griko
N B
NB
Skopinskaya
S N
SN
Yarkov
S P
SP
D000906:Antibodies; D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D001710:Biotin; D003165:Complement System Proteins; D004355:Drug Stability; D007118:Immunoassay / Q000379:methods; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D007202:Indicators and Reagents; D008081:Liposomes; D011506:Proteins / Q000032:analysis*; D013111:Spider Venoms / Q000032:analysis*; D019809:Streptavidin; D013997:Time Factors
D000906:Antibodies; D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D007202:Indicators and Reagents; D008081:Liposomes; D011506:Proteins; D013111:Spider Venoms; C020763:alpha-latrotoxin; D001710:Biotin; D003165:Complement System Proteins; D019809:Streptavidin