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</o:shapelayout></xml><![endif]--></head><body lang=EN-US link=blue vlink=purple><div class=WordSection1><p class=MsoNormal><span style='color:black'>Dear RASMB,<o:p></o:p></span></p><p class=MsoNormal><span style='color:black'><o:p> </o:p></span></p><p class=MsoNormal><span style='color:black'>We are trying to perform a measurement of v-bar using an Anton Paar DMA5000. The measurement is of a glycosylated protein. The densitometer was calibrated against a pure dI water standard and we’ve resorted to air checks between measurements to control for our cleaning protocol: 3 x detergent wash, 3 x dI water rinse, 3 x ethanol rinse, and air drying for 4 minutes or until the reading on the densitometer is 0.0012 g/cm3.<o:p></o:p></span></p><p class=MsoNormal><span style='color:black'><o:p> </o:p></span></p><p class=MsoNormal><span style='color:black'>We determined the extinction coefficient by synthetic boundary and believe we have accurately measured the concentration of our samples. Stock sample was dialyzed for a little over 48 hours against PBS and dilutions of the stock into dialysate were made in order to generate four solutions ranging from 0.1 to 0.3 mgs/mL.<o:p></o:p></span></p><p class=MsoNormal><span style='color:black'><o:p> </o:p></span></p><p class=MsoNormal><span style='color:black'>We are getting a values ranging between ~0.60 – 0.70, and are observing that the density measurements are not reproducible beyond the fourth decimal place. <o:p></o:p></span></p><p class=MsoNormal><span style='color:black'><o:p> </o:p></span></p><p class=MsoNormal><span style='color:black'>The reason for the lack of reproducibility appears to be that over time, while the sample is in the U-tube, small gas bubbles are observed inside the tube, either due to auto-hydrolysis of the solution or due to the small vibration of the tube or something else I am not aware of. We do not observe this for water alone. Samples were degassed, each for 10 minutes.<o:p></o:p></span></p><p class=MsoNormal><span style='color:black'><o:p> </o:p></span></p><p class=MsoNormal><span style='color:black'>My questions: <o:p></o:p></span></p><p class=MsoNormal><span style='color:black'>Has anyone observed similar issues when performing this measurement?<o:p></o:p></span></p><p class=MsoNormal><span style='color:black'>Is there an optimal buffer system for this measurement – perhaps at a salt concentration less than 100mM?<o:p></o:p></span></p><p class=MsoNormal><span style='color:black'>Is there, perhaps, a subtlety in this measurement that I need to be aware of?<o:p></o:p></span></p><p class=MsoNormal><span style='color:black'><o:p> </o:p></span></p><p class=MsoNormal><span style='color:black'>Thank you for any and all comments returned. They are greatly appreciated.<o:p></o:p></span></p><p class=MsoNormal><span style='color:black'><o:p> </o:p></span></p><p class=MsoNormal><span style='color:black'>Best regards<o:p></o:p></span></p><p class=MsoNormal><span style='color:black'>John Sumida<o:p></o:p></span></p><p class=MsoNormal><span style='color:black'>University of Washington</span><span style='font-size:12.0pt;font-family:"Courier New"'><o:p></o:p></span></p><p class=MsoNormal style='text-autospace:none'><span style='color:black'><o:p> </o:p></span></p></div></body></html>