ABSTRACT
LEDs present an alternative to lasers in LIF detection with CE, resulting in LED-induced fluorescence (LEDIF). LEDs are much less expensive, consume less energy and are more stable. In addition, LED light sources allow a greater range of wavelengths to better match the maximum wavelength for the fluorescence of the dye. Antibodies were largely studied in SDS capillary gel electrophoresis (SDS-CGE) and LIF detection with different dyes to label the proteins. In this work, our goal is to show that LEDs can advantageously replace lasers. We used 5-carboxytetramethylrhodamine succinimidyl ester (5-TAMRA.SE), 3-(2-furoyl)-quinoline-2 carboxaldehyde (FQ), and naphthalene-2,3-dialdehyde (NDA) to label IgG and we compared the LIF sensitivity with that obtained from LEDIF. We measured that the LOD values of LEDIF are identical to that obtained with the wavelength equivalent laser, and for 5-TAMRA.SE analysis, LOD values are about six times better than when the classical 488 nm laser was used.
Subject(s)
Electrophoresis, Capillary/instrumentation , Electrophoresis, Capillary/methods , Fluorescent Dyes/chemistry , Immunoglobulin G/analysis , Sodium Dodecyl Sulfate/chemistry , Spectrometry, Fluorescence/methods , Amines/chemistry , Amino Acids/chemistry , Fluorescein-5-isothiocyanate , Furans/chemistry , Humans , Immunoglobulin G/chemistry , Lasers , Limit of Detection , Naphthalenes/chemistry , Quinolines/chemistry , Rhodamines/chemistryABSTRACT
This paper describes the most important articles that have been published on amino acid analysis using CE during the period from June 2009 to May 2011 and follows the format of the previous articles of Smith (Electrophoresis 1999, 20, 3078-3083), Prata et al. (Electrophoresis 2001, 22, 4129-4138) and Poinsot et al. (Electrophoresis 2003, 24, 4047-4062; Electrophoresis 2006, 27, 176-194; Electrophoresis 2008, 29, 207-223; Electrophoresis 2010, 31, 105-121). We present new developments in amino acid analysis with CE, which are reported describing the use of lasers or light emitting diodes for fluorescence detection, conductimetry electrochemiluminescence detectors, mass spectrometry applications, and lab-on-a-chip applications using CE. In addition, we describe articles concerning clinical studies and neurochemical applications of these techniques.
Subject(s)
Amino Acids/analysis , Electrophoresis, Capillary , Animals , Humans , Mass Spectrometry , Spectrometry, Fluorescence , StereoisomerismABSTRACT
An analytical methodology for quality control analyses of IgG and their impurities is presented using a new UV-LIF (266 nm) detector inside the cassette of a CE instrument and its performance was evaluated. The observed sensitivity was very close to that obtained by silver staining of slab gels (LOD of 25 ng/mL), while the sensitivity of the analysis is 80 times better than with CE/UV absorption (214 nm). Examples of the analysis of pharmaceutical and other commercial IgGs are provided and the kinetics of the reduction of IgG by beta-mercaptoethanol is reported, demonstrating the ease of performing the analysis.
Subject(s)
Drug Contamination , Electrophoresis, Capillary/methods , Pharmaceutical Preparations/chemistry , Spectrometry, Fluorescence/methods , Animals , Immunoglobulin G/chemistry , Protein Denaturation , Rabbits , Rats , Sensitivity and Specificity , Silver StainingABSTRACT
This article describes the most important articles that have been published on amino acid analysis using CE during the period from June 2007 to May 2009. It follows the format of the previous articles of Smith [Electrophoresis 1999, 20, 3078-3083], Prata et al. [Electrophoresis 2001, 22, 4129-4138] and Poinsot et al. [Electrophoresis 2003, 24, 4047-4062; Electrophoresis 2006, 27, 176-194; Electrophoresis 2008, 29, 207-223]. For several years we have presented the new developments in amino acid analysis with CE which describe the use of laser emitting diodes for LIF as well as via MS. In addition, we describe articles concerning clinical studies and neuroclinical applications.
Subject(s)
Amino Acids/analysis , Electrophoresis, Capillary/methods , Agriculture , Animals , Biomedical Technology/methods , Body Fluids/chemistry , Humans , Lasers , Mass Spectrometry/methods , Neurotransmitter Agents/analysis , Spectrometry, Fluorescence/methods , StereoisomerismABSTRACT
An LIF detector was integrated into a CE system which uses a ball lens to focus the laser beam on the CE capillary. The detector employs an ellipsoid that is glued on the capillary window, to permit the collection of the fluorescence in the capillary. This 'trapped' fluorescence stays in the capillary because the angle of the silica/air interface is greater than the critical angle. The performance of this new detector setup is found to be identical to the collinear setup using the same ball lens. An application to the analysis of FITC-labeled IgG was optimized using a 14 cm effective length capillary. The LOD of an FITC-labeled IgG2 at an excitation wavelength of 488 nm was 150 pg/mL, which was 10 times better than the LOD recorded with slab gel silver staining. Using a tetramethylrhodamine (TAMRA)-labeled IgG2 and a 532 nm excitation wavelength the LOD is 50 pg/mL. The electropherograms of four different commercial FITC conjugates of IgG were studied. The presence of aggregates was observed in two samples while close kinetics of reduction was observed between free aggregates and high aggregates concentration samples. The integrated LIF detector provides an extremely powerful and convenient tool for antibody analysis and should be useful for therapeutic MAb control in pharmaceutical facilities.
Subject(s)
Electrophoresis, Capillary/instrumentation , Electrophoresis, Capillary/methods , Animals , Fluorescein-5-isothiocyanate , Fluorescence , Fluorescent Dyes , Humans , Immunoglobulin G/isolation & purification , Lasers , Mice , RabbitsABSTRACT
This paper describes a number of articles that have been published on amino acid (AA) analysis using CE during the period from June 2005 to May 2007. This review article follows the format of the previous articles of Smith (Electrophoresis 1999, 20, 3078-3083), Prata et al.. (Electrophoresis 2001, 22, 4129-4138), and Poinsot et al.. (Electrophoresis 2003, 24, 4047-4062 and Electrophoresis 2006, 27, 176-194). Several new developments in AA analysis with CE are reported describing the use of laser-emitting diodes for LIF, MS, and chips. In addition, we describe articles concerning clinical studies and neuroclinical applications.
Subject(s)
Amino Acids/analysis , Electrophoresis, Capillary/methods , Amino Acids/blood , Amino Acids/urine , Animals , Humans , Mice , RatsABSTRACT
This paper describes a number of articles that have been published on amino acid analysis using CE during the period from June 2003 to May 2005. This review article follows the previous ones of Smith (Electrophoresis 1999, 20, 3078-3083), Prata et al. (Electrophoresis 2001, 22, 4129-4138), and Poinsot et al. (Electrophoresis 2003, 24, 4047-4062). Several new developments in amino acid analysis with CE are reported concerning UV detection, LIF, MS, and NMR. In addition, we describe articles concerning clinical and pharmaceutical studies, neuroclinical applications, and agricultural and food analysis.
