[RASMB] Speed of Interference Optics vs. Absorbance Optics

[Lake Paul]

Folks,

I have a sample, a membrane protein with 0.01% DDM. I collected data at
45000 rpm, 4 min/scan. The sample is very heterogeneous but I am getting a
decent peak from the absorbance data (280 nm) at 7.4 S, but the interference
data gives me a peak at 6.5 S. I am trying to calculate the amount of
detergent bound so I need to align the interference data with absorbance
data. This is the first time in which I have gotten the peaks from the
interference data not lining up with absorbance data. Is this due to the
time it takes to make an absorbance scan relative to the interference scans?
And/or I just spun the sample too hard? Or is the DDM affecting the
sedimentation? See the attached powerpoint. 

Yes, the slit assembly is functioning properly J   

Any suggestions will be helpful! 

Best,

Lake

 

PS I collected data during the same run with the same sample except it had
GFP attached. The interference peak shifted accordingly. The DDM peak lined
up well also. 

 

Lake N. Paul, PhD

Biophysics Research Specialist

Discovery Park - Bindley Biosciences Center Purdue University

1203 West State Street

West Lafayette, Indiana, 47905

765.494.4960

BioAnalytical Lab Website:

http://www.purdue.edu/discoverypark/bioanalytical/

 

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