Question Regarding Scattering Intensity Differences After Peak Extraction in BIOXTAS RAW

[Siti Khadijah]

Hi, good morning. 

I am currently analyzing SEC-SAXS data and encountered a notable difference when using BIOXTAS RAW for peak extraction. Specifically, we observed that:

• Our SEC-SAXS chromatogram initially showed overlapping peaks, which we aimed to resolve to isolate the main elution peak.

• After applying peak extraction using BIOXTAS RAW, the scattering intensity profiles (I vs q) showed significantly higher absolute intensity compared to profiles processed without using BIOXTAS RAW.

• Although peak features were still visible without BIOXTAS RAW, we chose to use it to avoid ambiguity in the result and ensure more accurate frame selection.

Additionally, the I(0) value appears relatively high after extraction.

Attached are images showing the differences in scattering curves obtained with and without BIOXTAS RAW for your reference.

Could you kindly advise if such an increase in intensity is expected behavior following peak extraction? Or might this indicate an issue with baseline subtraction or scaling during the frame selection process?

Thank you very much for developing such a helpful tool and for your time and support.

Best regards,
Khadijah 

[Jesse Hopkins]

Hi Khadijah,

It would be useful to know how you generated the profile without RAW, and what you used to generate the profile with RAW. In RAW did you use a deconvolution approach like EFA or REGALS? If so, the deconvolved profiles are on an arbitrary scale, and so you wouldn't expect the intensity (or I(0)) to agree with profiles generated by direct averaging of the peak.

I would also strongly recommend that you trim the q range to the usable range (somewhere around 0.12?) before doing deconvolution, as all the noise below that point may influence the deconvolution.

All the best.

- Jesse

----

Jesse Hopkins, PhD (he/him)

Director, BioCAT

Sector 18, Advanced Photon Source

Research Associate Professor, Illinois Tech

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