It is difficult to raise antibodies against haptens and antigens that are unstable under the physiological conditions of the serum. Here we have used a phage antibody library to isolate antibody fragments against oxygen sensitive reduced flavin, by selection of the phage under anaerobic and reducing conditions at pH 5 and a pre-elution step with the oxidized flavin. The binding of the reduced hapten to one of the antibody fragments was characterised by time-resolved polarised fluorescence, and shown to be highly specific for the reduced flavin.
Bruggeman, Y. E., Boogert, A., Van Hoek, A., Jones, P. T., Winter, G., Schots, A., & Hilhorst, R. (1996). Phage antibodies against an unstable hapten: Oxygen sensitive reduced flavin. FEBS Letters, 388(2–3), 242–244. https://doi.org/10.1016/0014-5793(96)00544-3