Simple immunoassays for semiquantitative determination of calmodulin (CaM) and anti-CaM. Applications to affinity column chromatography of anti-CaM antibodies and estimation of CaM in synaptosomal membranes from rat cerebral cortex.
Dot-immunobinding assay for anti-calmodulin (CaM), and immunoblot assay for CaM have become feasible by proper choice of a fixative, i.e., formaldehyde vapor. The protocol for these simple and convenient immunological procedures is presented. Dot-immunobinding assay could be accomplished in 3 h, and immunoblot assay in 8 h. Activities of anti-CaM antibodies in fractions eluted from CaM affinity column were monitored by immunobinding assay. Semiquantitative immunoblot analysis was used to estimate CaM present in various fractions during preparation of EGTA-washed lysed synaptosomal membranes from rat cerebral cortex. CaM content in the final preparation was estimated at lower than 3 micrograms/mg of membrane protein.
Natsukari
N
N
Department of Psychiatry, Hamamatsu University School of Medicine, Japan.
Ohta
H
H
Fujita
M
M
D000818:Animals; D000906:Antibodies / Q000032:analysis*; D015153:Blotting, Western; D002147:Calmodulin / Q000032:analysis* / Q000276:immunology; D002540:Cerebral Cortex / Q000032:analysis*; D002846:Chromatography, Affinity; D004306:Dose-Response Relationship, Immunologic; D005404:Fixatives; D005557:Formaldehyde; D007118:Immunoassay; D015151:Immunoblotting; D066298:In Vitro Techniques; D051381:Rats; D013574:Synaptosomes / Q000032:analysis
D000906:Antibodies; D002147:Calmodulin; D005404:Fixatives; D005557:Formaldehyde