Haptoglobin Types in Dried Bloodstains

Abstract

THE determination of haptoglobin types in dried bloodstains has until recently met with very limited success. The method for determining haptoglobin types using starch gel^1–4 is the most satisfactory technique available for serum samples. Direct transposition of this technique to bloodstains has been tried by many research workers^5–7 both in starch and acrylamide gels. The patterns obtained from stain material differ considerably from those shown by fresh serum samples. It is extremely difficult to interpret the results as they can be obscured by the excess haemoglobin present and it can only be done if the stain is very fresh (that is, not more than about 3 days old). The problem of excess haemoglobin can be overcome by subjecting the sample to a short preliminary electrophoresis in starch gel at pH 6.5 (tank buffer: 0.04 M disodium hydrogen phosphate adjusted to pH 6.5 with ortho-phosphoric acid; gel buffer: 20 ml. tank buffer diluted to 150 ml.). At this pH, haemoglobin moves towards the cathode while the haptoglobin/haemoglobin complex moves towards the anode. After 30 min electrophoresis at 6 V/cm, a block of gel about 1 cm² is removed on the anodic side of the origin and inserted into a gel at pH 8.6 with a buffer system according to Poulik⁴. However, we required a method which was accurate and would allow the determination of haptoglobin types in stains at least one month old. These conditions have been met by using an immuno-electrophoresis method.