Capillary isoelectric focusing of proteins and microorganisms in dynamically modified fused silica with UV detection.
We suggest a method for the reproducible and efficient capillary isoelectric focusing of proteins and microorganisms in the pH gradient 3-10. The method involves the segmental injection of the simple ampholytes, the solution of the selected electrolytes, and the sample mixture of bioanalytes and carrier ampholytes to the fused silica capillaries dynamically modified by poly(ethylene glycol), PEG 4000, which is added to the catholyte, the anolyte and injected solutions. In order to receive the reproducible results, the capillaries were rinsed by the mixture of acetone/ethanol between analyses. For the tracing of the pH gradients the low-molecular-mass pI markers were used. The simple proteins and the mixed cultures of microorganisms, Saccharomyces cerevisiae CCM 8191, Escherichia coli CCM 3954, Candida albicans CCM 8180, Candida parapsilosis, Candida krusei, Staphylococcus aureus, Streptococcus agalactiae CCM 6187, Enterococcus faecalis CCM 4224, Staphylococcus epidermidis CCM 4418 and Stenotrophomonas maltophilia, were focused and separated by the method suggested. The minimum detectable number of microbial cells was 5x10(2) to 1x10(3) with on-column UV detection at 280 nm.
Horká
Marie
M
Institute of Analytical Chemistry, Academy of Sciences of the Czech Republic, Veverí 97, 611 42 Brno, Czech Republic. horka@iach.cz
Růzicka
Filip
F
Horký
Jaroslav
J
Holá
Veronika
V
Slais
Karel
K
D001419:Bacteria / Q000302:isolation & purification*; D002175:Candida / Q000302:isolation & purification; D013293:Enterococcus faecalis / Q000302:isolation & purification; D004926:Escherichia coli / Q000302:isolation & purification; D005658:Fungi / Q000302:isolation & purification*; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing / Q000295:instrumentation / Q000379:methods*; D011092:Polyethylene Glycols / Q000737:chemistry; D011506:Proteins / Q000032:analysis* / Q000302:isolation & purification; D015203:Reproducibility of Results; D012441:Saccharomyces cerevisiae / Q000302:isolation & purification; D012822:Silicon Dioxide / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet / Q000379:methods; D013211:Staphylococcus aureus / Q000302:isolation & purification; D013212:Staphylococcus epidermidis / Q000302:isolation & purification; D020615:Stenotrophomonas maltophilia / Q000302:isolation & purification; D013292:Streptococcus agalactiae / Q000302:isolation & purification
D011506:Proteins; D011092:Polyethylene Glycols; D012822:Silicon Dioxide