Concentration by centrifugation for gas exchange EPR oximetry measurements with loop-gap resonators.
Measurement of the bimolecular collision rate between a spin label and oxygen is conveniently carried out using a gas permeable plastic sample tube of small diameter that fits a loop-gap resonator. It is often desirable to concentrate the sample by centrifugation in order to improve the signal-to-noise ratio (SNR), but the deformable nature of small plastic sample tubes presents technical problems. Solutions to these problems are described. Two geometries were considered: (i) a methylpentene polymer, TPX, from Mitsui Chemicals, at X-band and (ii) Teflon tubing with 0.075 mm wall thickness at Q-band. Sample holders were fabricated from Delrin that fit the Eppendorf microcentrifuge tubes and support the sample capillaries. For TPX, pressure of the sealant at the end of the sample tube against the Delrin sample holder provided an adequate seal. For Teflon, the holder permitted introduction of water around the tube in order to equalize pressures across the sealant during centrifugation. Typically, the SNR was improved by a factor of five to eight. Oxygen accessibility applications in site-directed spin labeling studies are discussed.
Subczynski
Witold K
WK
Department of Biophysics, Medical College of Wisconsin, Milwaukee, WI 53226, USA.
Felix
Christopher C
CC
Klug
Candice S
CS
Hyde
James S
JS
D002498:Centrifugation / Q000295:instrumentation* / Q000379:methods; D004578:Electron Spin Resonance Spectroscopy / Q000295:instrumentation* / Q000379:methods; D004867:Equipment Design; D019544:Equipment Failure Analysis; D017022:Flow Injection Analysis / Q000295:instrumentation* / Q000379:methods; D005740:Gases / Q000032:analysis*; D044085:Microfluidics / Q000295:instrumentation / Q000379:methods; D010092:Oximetry / Q000295:instrumentation* / Q000379:methods; D010100:Oxygen / Q000032:analysis*; D014159:Transducers
D005740:Gases; D010100:Oxygen