ORIGINAL PAPER A high-efficiency sample introduction system for capillary electrophoresis analysis of amino acids from dynamic samples and static dialyzed human vitreous samples Eric E. Patterson II & Sujeewa C. Piyankarage & Kyaw ThetMaw Myasein & Jose S. Pulido & Robert F. Dundervill III & R. Mark Hatfield & Scott A. Shippy Received: 15 May 2008 / Revised: 8 July 2008 / Accepted: 17 July 2008 / Published online: 12 August 2008 # Springer-Verlag 2008 Abstract A low-volume automated injection system for the analysis of chemically complex, amino acid samples is presented. This system utilizes submicroliter sample vol- umes stored on a 75-μm inner diameter capillary. A pulse of positive pressure (82 kPa) is used to load nanoliter sample volumes into an in-house fabricated interface and onto a separation capillary. Residual sample solution in the inter- face is immediately washed away by a continuous transverse flow through the injection interface, yielding a sharp and reproducible sample plug. By performing multiple injections of a static sample, one may average the signals to yield a signal-to-noise ratio improvement of up to 4.07-fold for 20 injections compared with a theoretical maximum of a 4.47-fold improvement. Without interruption of the applied voltage, injections performed every 150 s were used to monitor the progress of the reaction of multiple amino acids with the fluorogenic dye 3-(4-carboxybenzoyl)quinoline-2- carboxaldehyde. Analysis of dialyzed clinical vitreous sam- ples demonstrates the resolution and quantitation of arginine, lysine, leucine, glutamine, and glutamate. Observed levels are comparable with those of nonautomated injection methods and reports by others. Keywords Signal averaging . Capillary electrophoresis . Injection . Sample introduction . Dialysis . Vitreous Introduction Amino acid analysis is important to the understanding of mammalian physiological function, and capillary electro- phoresis (CE) is quickly becoming the customary analysis method for these volume-limited biological samples [1–5]. This is due to the fast separation capability, low sample volume requirements, and high separation resolution of CE. With primary amine reactive dyes, the sensitivity of CE for amino acid analysis becomes very high and the detection becomes very selective [6]. The need to analyze amino acids in biological samples is clear from their potential role either as precursors to or products of a diseased state [7–10]. Efficient and sensitive amino acid analysis, with minimal sample volume requirements, can elucidate normal physio- logical function and highlight dysregulation due to disease. A prime example of the need for further development of analytical methods for amino acid analysis is seen with the increasing consensus that excitatory amino acid levels are elevated with retinal disease [7–11]. Better sample collec- tion and more efficient analysis methods for these samples will be critical in elucidating an understanding of these problems. To fully utilize the small-volume advantages that CE offers, efficient methods of sample injection must be developed. With use of traditional sample introduction methods, a very small Anal Bioanal Chem (2008) 392:409–416 DOI 10.1007/s00216-008-2304-5 E. E. Patterson II : S. C. Piyankarage : K. T. Myasein : S. A. Shippy (*) Department of Chemistry, M/C 111, University of Illinois at Chicago, Chicago, IL 60607, USA e-mail: sshippy@uic.edu J. S. Pulido Department of Ophthalmology, Mayo Clinic, Rochester, MN 55905, USA R. F. Dundervill III : R. M. Hatfield Retina Consultants, Charleston, WV 25301, USA