Attomole sensitivity for unlabeled proteins and polypeptides with on-chip capillary electrophoresis and universal detection by interferometric backscatter

• Published in: Electrophoresis 2003-03, Vol.24 (5), p.865-873
• Main Authors: Wang, Zhanling, Swinney, Kelly, Bornhop, Darryl J.
• Format: Article
• Language: English
• Subjects: Animals; Backscattering interferometry; Buffers; Calmodulin - analysis; Cattle; Egg Proteins - analysis; Electrophoresis, Capillary - methods; Interferometry - instrumentation; Label-free protein quantification; Lactalbumin - analysis; Microchemistry - instrumentation; On-chip capillary electrophoresis; Pepsinogen A - analysis; Peptides - analysis; Peptides - chemistry; Protein Denaturation; Proteins - analysis; Proteins - chemistry; Refractive index detection; Sensitivity and Specificity; Serum Albumin, Bovine - analysis; Sodium Dodecyl Sulfate
• ISSN: 0173-0835; 1522-2683
• DOI: 10.1002/elps.200390109

A universal detector based on backscatter interferometry has been developed to perform nanoliter volume refractive index measurements for on‐chip sodium dodecyl sulfate (SDS) gel based (polyethylene oxide gel) separations and quantification label‐free proteins. The on‐chip interferometric backscatter detector (OCIBD) system consists of a simple, folded optical train based on the interaction of a laser beam with an etched channel in the shape of half cylinder in a fused‐silica plate. The backscattered light from the channel takes on the form of a high‐contrast interference pattern that contains information related to the bulk properties of the fluid located within the probe or detection volume of 2.32×10−9 L. Depending on capillary electrophoresis (CE) injection method, the positional changes of the interference pattern extrema (fringes) allow for the quantification of unlabeled proteins at levels ranging from 11 to 310 amol (2.7×10−8 mol/L) with a linear dynamic range of 2.5 decades (egg albumin). Using OCIBD microchannel‐based SDS capillary gel electrophoresis (SDS/CGE), separation and detection of five label‐free proteins was achieved in less than 100 seconds with detection limits ranging from 0.95 pg (1.1×10−16 mol or 2.5×10−7 mol/L) of calmodulin to 7.0 pg (1.0×10−16 mol or 2.4×10−7 mol/L) for bovine serum albumin (BSA) without signal filtering or active thermal control. This development shows that a universal detector based on backscatter interferometry can be used effectively for on‐chip label‐free solute analysis.