2.5 ǀ Expression of cpOAS1 protein using western blotting
Caprine OAS1 proteins expressed in goat endometrial cells were separated on a 12% SDS-PAGE gel and transferred electrophoretically to a PVDF membrane (BiotraceTM PVDF, PALL Corporation, India) at 100 V for 2 h uteri as described earlier58. After transfer, the membrane was incubated in blocking buffer [5% (w/v) skim milk powder in PBS, pH 7.4] at room temperature for 1h with gentle agitation. To detect the cpOAS1 isoform proteins expressed in the goat endometrium, the membranes were incubated with rabbit OAS1 polyclonal antibody (Invitrogen, Catalog # PA5-41755) in 1:800 dilutions in blocking buffer at 4˚C overnight. To detect overexpressed cpOAS1 isoforms, the membranes were incubated at room temperature for 1.5 h with HRP conjugated mouse anti-rabbit secondary antibodies (Santa Cruz, CA, USA) to a 1:2000 dilution of in PBS. DAB substrate buffer system (GeNei, India) was used to detect the presence of secondary antibody. The densitometric analysis of protein was performed by NIH ImageJ 1.44p software (National Institutes of Health, Bethesda, Maryland, USA). The relative intensity of the bands was quantified and normalized with β-actin (Santa Cruz, CA, USA).