An Improved Microprocedure for the Determination of Heparin

The microelectrophoresis method described by Jaques et al. (Can. J. Physiol. Pharmacol. 46, 351–360 (1968)) for identification and quantitation of microquantities of heparin was improved by using the differential effects of inorganic electrolytes on the solubility of quaternary ammonium – mucopolysaccharide complex. Samples were applied to agarose gel coated microscope slides for electrophoresis. The slides were then immersed in 0.1% cetylpyridinium chloride (CPC) in M NaCl at 35 °C with constant agitation for 5 h, washed thoroughly with 0.1% CPC, then stained with toluidine blue. As a result of this treatment, only heparin in concentrations of 0.25–2.0 μg produced optically dense spots on the slides for measurement. Heparin was identified by the combined information from electrophoretic migration, salt fractionation, and metachromatic activity.