Bio-Rad Immun-Star™ AP Chemiluminescence Kits - Manual (Page 18)

2.4 Detailed Assay Procedure
Note: Before beginning, read through the entire procedure.
1. Antigen application – apply antigen to the membrane
surface using one of the three basic methods described
below. A small amount of known antigen or primary
antibody dotted on one corner of the membrane prior to
blocking will produce a positive reaction if the procedure is
successful.
a. Electrophoretic blotting – the antigens of interest
are electrophoretically transferred to the membrane from
a gel (i.e., SDS-PAGE gel, IEF gel, or native gel) using
the Trans-Blot
® cell, Mini Trans-Blot ® cell, Criterion ®
blotter, or Trans-Blot SD cell. If desired, cut the wet
membrane into 0.6–0.8 cm wide strips. Immerse the
strips or the entire sheet in TBS before proceeding to
the blocking step.
b. Microfiltration blotting – the Immun-Star assay can
easily be adapted for use in the Bio-Dot ® and Bio-Dot
SF apparatus. These instruments allow rapid,
reproducible applications of up to 96 samples on one
membrane sheet. The membrane should be removed
from the apparatus after antigen application. Because
nonfat dry milk cannot be filtered through the
membrane in the Bio-Dot or Bio-Dot SF apparatus, the
blocking and incubation steps should be carried out in
a separate container.
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