Conjugated antibodies are a useful research tool in a variety of applications, ranging from Western blot to flow cytometry. We at Novus Biologicals offer many primary antibodies conjugated to enzymes such as HRP, fluorochromes such as FITC, and others including biotin and cyanine dyes.
Our antibody lab is always busy with custom antibody conjugation projects. This past week the lab conducted numerous conjugations, including a LDL Receptor antibody to biotin, a HMGB1 antibody to HRP, an Actin antibody to DyLight 488, and a CD133 antibody to DyLight 549.
When asked about the difference between conjugating an antibody to HRP, biotin and DyLight dyes, one of the Novus lab technicians, David Kuroki, explained just one of the variances:
“Dialysis is used to remove primary amines and any sodium azide that may be present. For HRP and Biotin the dialysis buffer is PBS, pH 7.4. For the DyLight fluors the dialysis buffer is 50 mM Sodium Borate, pH 8.5. There is also a difference in incubation times, 30 minutes for Biotin, 1 hour for DyLight fluors and 3 hours for HRP.”
Kuroki also mentioned that antibody conjugations can take anywhere from 3 to 6 hours, depending on the specific conjugate. First the antibody is quantitated for a reference point, then dialyzed into the appropriate buffer. Next the antibody is concentrated, if necessary, and then quantitated again to determine recovery efficiency. Following the completion of these steps, the actual conjugation takes place.
Browse the conjugated antibodies that we offer by visiting our primary antibody page and using the conjugate filter on the bottom, left hand side of the screen. In addition, easily label any antibody with only 30-seconds hands-o time using our Lightning-Link Antibody Labeling Kits.