Processing and analysis of SEC-SAXS data Alejandro Sasha Panjkovich - - PowerPoint PPT Presentation

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Processing and analysis of SEC-SAXS data Alejandro Sasha Panjkovich - - PowerPoint PPT Presentation

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Processing and analysis of SEC-SAXS data Alejandro Sasha Panjkovich Celgene Corporation 2019.10.19 A. Panjkovich (Celgene) SEC-SAXS 2019.10.19 1 / 7 Outline Visualize elution profile in terms of scattering Common experimental issues

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SLIDE 1

Processing and analysis of SEC-SAXS data

Alejandro ‘Sasha’ Panjkovich

Celgene Corporation

2019.10.19

  • A. Panjkovich (Celgene)

SEC-SAXS 2019.10.19 1 / 7

slide-2
SLIDE 2

Outline

Visualize elution profile in terms

  • f scattering

Common experimental issues Identify sample and buffer region Evaluate sample behaviour (Rg and MW) Align with additional data

  • A. Panjkovich (Celgene)

SEC-SAXS 2019.10.19 2 / 7

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SLIDE 3

CHROMIXS: CHROMatography Inline X-ray Scattering

Visualize SEC-SAXS run Manual or automatic region selection Compare additional parameters and data Export data and figures Complete automatic processing

average intensity

8,500 9,000 9,500 10,000 10,500 11,000 11,500

radius of gyration

1 2 3 4 5 6

frame nr.

400 500 600 700 800 900 BSA SEC-SAXS monomer peak buffer Rg values

CHROMIXS

dimer peak
  • A. Panjkovich (Celgene)

SEC-SAXS 2019.10.19 3 / 7

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SLIDE 4

Experimental issues

  • A. Panjkovich (Celgene)

SEC-SAXS 2019.10.19 4 / 7

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SLIDE 5

Experimental issues

  • A. Panjkovich (Celgene)

SEC-SAXS 2019.10.19 5 / 7

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SLIDE 6

Reference and other programs for further analysis

CHROMIXS (part of ATSAS package) Panjkovich and Svergun (2018) CHROMIXS: automatic and interactive analysis of chromatography-coupled small angle X-ray scattering data. Bioinformatics 34:1944-46 US SOMO Brookes et al. (2016) ”US-SOMO HPLC-SAXS Module: Dealing with Capillary Fouling, and Extraction of Pure Component Patterns from Poorly Resolved SEC-SAXS Data”, J. Appl. Cryst. 49. BioXTAS RAW Hopkins, J. B., Gillilan, R. E., and Skou, S. (2017). Bioxtas raw: improvements to a free open-source program for small-angle x-ray scattering data reduction and analysis. Journal of applied crystallography, 50, 1545–1553. Scatter http://www.esrf.eu/UsersAndScience/Experiments/CRG/BM26/SaxsWaxs/DataAnalysis/Scatter

  • A. Panjkovich (Celgene)

SEC-SAXS 2019.10.19 6 / 7

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SLIDE 7

Summary

Visualize elution profile in terms

  • f scattering

Common experimental issues Identify sample and buffer region Evaluate sample behaviour (Rg and MW) Align with additional data

  • A. Panjkovich (Celgene)

SEC-SAXS 2019.10.19 7 / 7

slide-8
SLIDE 8

Structural and biophysical methods for biological macromolecules in solution

EMBO Global Exchange Lecture Course 14 – 20 October 2019 | Santiago, Chile