...n Tris, cannot have any free primary amines other than protein, we dialyze protein into PBS before labelling)
*Add 350uL of PBS and 25ug of protein to an eppie.
...
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#Calculate extinction coefficient for protein of interest
...] tool to calculate the extinction coefficient in both molar and mg/mL for protein. Assume all Cys residues are half-cysteines.
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702 bytes (104 words) - 14:03, 11 May 2009
*BioRad Protein Assay Dye Reagent Concentrate cat#500-0006
#Add 1-10 uL of protein sample, cover with parafilm and mix
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1 KB (202 words) - 14:39, 15 March 2018
! Protein
[[Category:Protein Purification]]
...
444 bytes (57 words) - 17:54, 24 July 2012
* combine 0.5 ml lysate, 0.2-0.5 ug of antibody and 25 ul of protein A/G plus agarose beads (catalog # sc 2003 santa cruz).
* for tissue lysates - use 0.5 mg of protein, add primary antibody first for 0.5 hour and then add beads.
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1 KB (237 words) - 14:23, 21 August 2016
*Express and induce protein in culture under appropriate conditions:
...ncentration of IPTG and duration of induction should be optimized for each protein)
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2 KB (337 words) - 14:40, 12 August 2009
*Express and induce protein in culture under appropriate conditions:
...ncentration of IPTG and duration of induction should be optimized for each protein)
...
2 KB (340 words) - 22:00, 2 December 2010
Useful for blocking protein synthesis
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156 bytes (23 words) - 12:55, 11 July 2014
[[ Category: Protein Purification ]]
[[ Category: Protein-Protein Interactions ]]
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3 KB (572 words) - 17:28, 31 July 2012
*Protein of interest (free of DTT/Glycerol/Glutathione)
#Prepare assay volumes of 50 uL in TLA-100 tubes with ~10 uM protein and varying lipid concentrations (for a first try use 0-1 mM lipid at 10X d
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2 KB (260 words) - 13:01, 2 November 2010
#Check that the HPLC is set up for protein work and not radioactive work (see [[HPLC - Plumbing Setup]]).
##Place the protein lead into the equillibration buffer and prime.
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3 KB (559 words) - 19:47, 9 February 2011
#Follow Protein Lysate instructions for Bradford Assay (see [[Bradford Assay]])
...2X loading buffer with B-ME to each lyaste. This will generate a 2 mg/mL protein solution in SDS Loading Buffer
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1 KB (253 words) - 16:06, 25 April 2019
* Protein of interest to be immobilized. Ideally start with 10-100 ug/mL protein (need ~100 uL per surface)
...to start). Lower pH will give better immobilization, but potentially less protein function
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2 KB (302 words) - 14:43, 16 June 2010
===Protein Analysis===
*[[Surface Plasmon Resonance - Protein Lipid Interactions]]
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4 KB (484 words) - 12:10, 15 August 2016
...e first Refseq mRNA (should start with NM) then click on that and find the protein (should start with NP)
...gov/Blast.cgi?PROGRAM=blastp&PAGE_TYPE=BlastSearch&LINK_LOC=blasthome NCBI Protein Blast].
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3 KB (360 words) - 13:16, 18 April 2019
*Express and induce protein in culture under appropriate conditions:
...ncentration of IPTG and duration of induction should be optimized for each protein)
...
3 KB (440 words) - 17:55, 24 July 2012
Protein must be extensively dialysed in PBS to remove any amine group from the samp
#Dissolve protein ligand (~1mg) in 6 mL of coupling buffer.
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2 KB (409 words) - 00:15, 27 May 2009
[[ Category: Protein Purification ]]
[[ Category: Protein-Protein Interactions ]]
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2 KB (355 words) - 17:46, 31 July 2012
*Protein Markers, we use SeaBlue Plus2 from Invitrogen (cat #LC5925)
...comb out and rinse with water. Pick a percent gel based on your expected protein size.
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3 KB (477 words) - 17:46, 20 January 2025
# Measure protein content by bradford assay for normalization
...ter glycosidase treatment and are presented in equivalent glucose units/mg protein.
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1 KB (173 words) - 18:23, 30 June 2022