Raising the Dead Volume: Analysis of Microsamples Diluted and Corrected with Near Infrared Tracer

Sample processing robotics require large liquid volumes to operate efficiently. Robotics are impractical in settings that deal in small specimen volumes such as pediatric laboratories. Short of manual sample handling, remedies for the current state include a redesign of current hardware or specialized adaptation for submilliliter specimens.

We blindly increased the volume of plasma specimens with diluent containing a near infrared dye, IR820, to assess the change to the original specimen volume. Diluted specimens were analyzed using a variety of assay formats/wavelengths (sodium, calcium, alanine aminotransferase, creatine kinase, cholesterol, HDL cholesterol, triglyceride, glucose, total protein, creatinine), and results were compared to neat specimens. Recovery of analyte in the diluted specimens vs neat was the primary outcome measure.

Mean analytic recovery from the diluted specimens across all assays ranged from 93% to 110% after correction using IR820 absorbance. Absorbance correction compared favorably to mathematical correction using known volumes of specimens and diluents (93%-107%). Pooled mean analytic imprecision across all assays ranged from 2% using the neat specimen pool to 8% when plasma pool was diluted to 30% of its original concentration. No interference from dye addition was noted, indicating the diluent was broadly applicable and chemically inert. The greatest variability in recovery was observed when respective analyte concentrations were present near the lower limits of assay detectability.

Addition of a chemically inert diluent containing a near-infrared tracer is a feasible way to raise specimen dead volume and potentially automate processing and measurement of clinical analytes in microsamples.