Alkaline Phosphatase-Immunoglobulin Conjugate Glutaraldehyde Protocol

9 Alkaline Phosphatase-Immunoglobulin Conjugate (Glutaraldehyde Protocol) 

The advantage of the use of alkaline phosphatase (AP) lies in the high purity of commercially available enzyme. But it is difficult to get such highly specific activities as for HRP, because there is no simple procedure for separation of unbound antibodies from the conjugate. 

15 rrd Soln. A to the dialysate. Agitate at RT for 2 h and continue at 4 °C overnight. Dialyze three times against 100 ml PBS for 3 h each. Bring the dialysate up to 2 mg BSA and fill up to 10 ml with Soln. B. Aliquote and store at -20 °C. 

A separation of conjugated AP from unbound enzyme in EIA is not necessary, because free AP is removed during the washing steps. 

1 Enzymatic Reaction of Aikaiine Phosphatase from Caif intestine 

---