May 24, 2022

Public workspaceProtein extraction protocol

DOI
https://dx.doi.org/10.17504/protocols.io.5jyl896o6v2w/v1
  • rabdelha 1
  • 1California Institute of Technology
rabdelha
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DOI: https://dx.doi.org/10.17504/protocols.io.5jyl896o6v2w/v1
External link: https://doi.org/10.7554/eLife.81453
Protocol Citation: rabdelha 2022. Protein extraction protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.5jyl896o6v2w/v1
Manuscript citation:
Abdel-Haq R, Schlachetzki JC, Boktor JC, Cantu-Jungles TM, Thron T, Zhang M, Bostick JW, Khazaei T, Chilakala S, Morais LH, Humphrey G, Keshavarzian A, Katz JE, Thomson M, Knight R, Gradinaru V, Hamaker BR, Glass CK, Mazmanian SK, A prebiotic diet modulates microglial states and motor deficits in α-synuclein overexpressing mice. eLife doi: 10.7554/eLife.81453
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it’s working
Created: May 16, 2022
Last Modified: May 31, 2024
Protocol Integer ID: 62652
Keywords: Protein extraction, protease inhibitor, Tissue extraction, ASAPCRN, protocol details protein extraction procedure, protein extraction protocol, extraction, protocol detail, protocol this protocol, protein, protocol, procedure
Abstract
This protocol details protein extraction procedure.
Attachments
Icon for 440-928.docx
440-928.docx
13KB
Materials

ReagentTissue Extraction ReagentThermo Fisher ScientificCatalog #FNN0071 :

  • Warm reagent to TemperatureRoom temperature .
  • Add protease inhibitor tablet (1 tablet/Amount10 mL ).

ABC
Weight of SN from 1 side5 mgAdd 400 µL reagent
Weight of STR from 1 side15 mgAdd 400 µL reagent
Weight of CB from 1 side40 mgAdd 400 µL reagent
Weight of cortex from 1 side30 mgAdd 300 µL reagent









Troubleshooting
Protein extraction protocol
5m
Warm reagent to TemperatureRoom temperature .

Add protease inhibitor tablet (1 tablet/Amount10 mL ) (complete EDTA free).

Pipetting
Add amount of reagent to each sample and transfer to bead beating tube.
Pipetting
Homogenize tissue sample in bead beater (see protocol in notebook), speed 3; let samples sit TemperatureOn ice for 10-Duration00:15:00 after to settle.
15m
Centrifuge homogenized sample at Centrifigation10000 rpm for Duration00:05:00 .

5m
Centrifigation
Collect supernatants and store in Temperature-80 °C .