Analysis of a biopolymer by capillary electrophoresis with a chemiluminescence detector using a polymer solution as the separation medium.
We developed capillary electrophoresis with a chemiluminescence detector using a polymer solution as the separation medium for the analysis of biopolymers, such as DNA and protein. A peroxyoxalate chemiluminescence reagent of bis(2,4,6-trichlorophenyl)oxalate was used together with fluorescein-labeling reagent. When a migration buffer solution containing carboxylmethylcellulose was used, the flow-type chemiluminescence detection cell was found to give a better resolution than the batch-type one. Fluorescein-labeled adenosine triphosphate of 1.0 x 10(-4) M was examined by means of capillary electrophoresis with absorption (260 nm), fluorescence (ex. 496 nm and em. 517 nm), and chemiluminescence detectors. The chemiluminescence detection showed the highest sensitivity among them; the S/N ratios obtained by absorption, fluorescence, and chemiluminescence detections were 4, 38, and 130, respectively. Fluorescein-labeled DNA was prepared through a polymerase chain reaction using fluorescein-labeled deoxyadenosine triphosphate. A mixture of the labeled DNA fragments (500, 600, 700, 800, 900, and 993 bp) was successfully separated and detected by the present system. A mixture of proteins (lysozyme, cytochrome C, and ribonuclease A) which were labeled with fluorescein isothiocyanate was also separated and detected.
Tsukagoshi
Kazuhiko
K
Department of Chemical Engineering and Materials Science, Faculty of Engineering, Doshisha University, Kyotanabe, Kyoto 610-0321, Japan.
Shikata
Yukihiro
Y
Nakajima
Riichiro
R
Murata
Masaharu
M
Maeda
Mizuo
M
D000042:Absorption; D000255:Adenosine Triphosphate / Q000032:analysis; D020029:Base Pairing; D001704:Biopolymers / Q000032:analysis* / Q000737:chemistry*; D002021:Buffers; D003574:Cytochrome c Group / Q000032:analysis / Q000737:chemistry; D004247:DNA / Q000032:analysis / Q000737:chemistry; D019075:Electrophoresis, Capillary / Q000379:methods*; D019793:Fluorescein; D008163:Luminescent Measurements; D009113:Muramidase / Q000032:analysis / Q000737:chemistry; D010070:Oxalates; D011108:Polymers / Q000737:chemistry*; D015203:Reproducibility of Results; D012259:Ribonuclease, Pancreatic / Q000032:analysis / Q000737:chemistry; D012680:Sensitivity and Specificity; D012996:Solutions / Q000737:chemistry; D013997:Time Factors
D001704:Biopolymers; D002021:Buffers; D003574:Cytochrome c Group; D010070:Oxalates; D011108:Polymers; D012996:Solutions; C036521:bis(2,4,6-trichlorophenyl)oxalate; D000255:Adenosine Triphosphate; D004247:DNA; D012259:Ribonuclease, Pancreatic; D009113:Muramidase; D019793:Fluorescein