A microtechnique for dialysis of small volume solutions with quantitative recoveries.
An improved microtechnique designed for dialysis of solutions with volumes ranging from less than 10 microliter up to approximately 600 microliter is described. Samples, dispensed in Microfuge tubes, are dialyzed in situ across dialysis membrane secured over the tube opening by a perforated Microfuge tube cap. The retentate is efficiently recovered by centrifugation at 10,000g for 10 s. Fifty percent escape (E50) times of [14C]glycine from 25-microliter solutions of soybean trypsin inhibitor (0.1, 1.0, 4.0 mg/ml) in 0.2 M NaCl were approximately 19 min. The E50 times of 3H2O increased in a linear fashion from 2.7 min for 25-microliter samples to 75 min for 600-microliter samples of H2O (pH 7.0, 4 degrees C). The mean permeability coefficient (P) of the dialysis membrane to 3H2O during microdialysis, calculated as 3.0 X 10(-4) cm/s, was similar to membrane permeability coefficients reported for dialysis by conventional methods. Quantitative recoveries (greater than approximately 90%) of [14C]glycine-labeled type I collagen, [methyl-14C]antithrombin III, and [32P]DNA were achieved after microdialysis.
Medical Research Council Group in Periodontal Physiology, University of Toronto, Ontario, Canada.
C M
CM
Overall
D000818:Animals; D004247:DNA / Q000302:isolation & purification; D003956:Dialysis / Q000295:instrumentation*; D005998:Glycine / Q000302:isolation & purification; D007202:Indicators and Reagents; D008832:Microchemistry; D010539:Permeability; D051381:Rats
D007202:Indicators and Reagents; D004247:DNA; D005998:Glycine