Simple, miniaturized blood plasma extraction method.

Abstract

A rapid plasma extraction technology that collects a 2.5 μL aliquot of plasma within three minutes from a finger-stick derived drop of blood was evaluated. The utility of the plasma extraction cards used was that a paper collection disc bearing plasma was produced that could be air-dried in fifteen minutes and placed in a mailing envelop for transport to an analytical laboratory. This circumvents the need for venipuncture and blood collection in specialized vials by a phlebotomist along with centrifugation and refrigerated storage. Plasma extraction was achieved by applying a blood drop to a membrane stack through which plasma was drawn by capillary action. During the course of plasma migration to a collection disc at the bottom of the membrane stack blood cells were removed by a combination of adsorption and filtration. After the collection disc filled with an aliquot of plasma the upper membranes were stripped from the collection card and the collection disc was air-dried. Intercard differences in the volume of plasma collected varied approximately 1% while volume variations of less than 2% were seen with hematocrit levels ranging from 20% to 71%. Dried samples bearing metabolites and proteins were then extracted from the disc and analyzed. 25-Hydroxy vitamin D was quantified by LC-MS/MS analysis following derivatization with a secosteroid signal enhancing tag that imparted a permanent positive charge to the vitamin and reduced the limit of quantification (LOQ) to 1 pg of collected vitamin on the disc; comparable to values observed with liquid-liquid extraction (LLE) of a venipuncture sample. A similar study using conventional proteomics methods and spectral counting for quantification was conducted with yeast enolase added to serum as an internal standard. The LOQ with extracted serum samples for enolase was 1 μM, linear from 1 to 40 μM, the highest concentration examined. In all respects protein quantification with extracted serum samples was comparable to that observed with serum samples obtained by venipuncture.