Difference between revisions of "Western Blot of bovine casein"
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'''Material:''' for SDS page: | '''Material:''' for SDS page: | ||
SDS (10%) catalog number: SDS10-100 | |||
SDS (10%) catalog number: SDS10-100 | |||
distilled water | distilled water | ||
Tris-Glycine SDS Running Buffer 10X stock (Novex Life Tech, Catalog number: LC2675) | Tris-Glycine SDS Running Buffer 10X stock (Novex Life Tech, Catalog number: LC2675) | ||
Tris-Glycine Mini Gels 4-20% 1.0mm x 10 wells/gel, (Catalog number: EC6025Box, Lot 14082110, received September 2014, possibly stored at -20C for a period of time) | Tris-Glycine Mini Gels 4-20% 1.0mm x 10 wells/gel, (Catalog number: EC6025Box, Lot 14082110, received September 2014, possibly stored at -20C for a period of time) | ||
Protein ladder (10-250 kDa) (NEB, Catalog number: p7703S) | Protein ladder (10-250 kDa) (NEB, Catalog number: p7703S) | ||
6X loading dye + SDS solution (Fermentas) | 6X loading dye + SDS solution (Fermentas) | ||
Micellular casein powder from CCL stock (for positive control) | Micellular casein powder from CCL stock (for positive control) | ||
Gel box and power supply from BioCurious | |||
Simply Blue Safe Stain LC6060 Lot#1540663 | |||
'''Results:''' | '''Results:''' | ||
Preparation of casein control | |||
Take 500ul of distilled water in eppendorf tube + few grains of casein powder (use 200ul hp). Vortex vigorously. | |||
Preparation of samples to load on gel | |||
Take 40ul of 1, 8, 3B, 6B, +7F also the negative control to eppendorf tubes | |||
5ul of 6x loading dye to each tube (this isn't to final conc. of 1x; should be 8ul) | |||
Boil all the tubes for 10 mins at 95C (in order to denature the protein structure to linear) | |||
Loading & Running Gel Generally, followed manufacturer's protocol for running Tris-Glycine 4%-20% mini-gels, Media:LifeTech_NuPAGE_TrisGlycine_Mini_Gels_Cat-EC6025BOX.pdf | |||
Latest revision as of 01:06, 3 May 2015
May 2, 2015: Western Blot of Bovine Casein
Location: BioCurious
Participants: Priyanka, Johan, Advait, Maria, Patrik
Notes: Priyanka, Johan, Advait
Aims: Confirm presence of bovine casein protein
Material: for SDS page:
SDS (10%) catalog number: SDS10-100
distilled water
Tris-Glycine SDS Running Buffer 10X stock (Novex Life Tech, Catalog number: LC2675)
Tris-Glycine Mini Gels 4-20% 1.0mm x 10 wells/gel, (Catalog number: EC6025Box, Lot 14082110, received September 2014, possibly stored at -20C for a period of time)
Protein ladder (10-250 kDa) (NEB, Catalog number: p7703S)
6X loading dye + SDS solution (Fermentas)
Micellular casein powder from CCL stock (for positive control) Gel box and power supply from BioCurious
Simply Blue Safe Stain LC6060 Lot#1540663
Results:
Preparation of casein control
Take 500ul of distilled water in eppendorf tube + few grains of casein powder (use 200ul hp). Vortex vigorously.
Preparation of samples to load on gel
Take 40ul of 1, 8, 3B, 6B, +7F also the negative control to eppendorf tubes
5ul of 6x loading dye to each tube (this isn't to final conc. of 1x; should be 8ul)
Boil all the tubes for 10 mins at 95C (in order to denature the protein structure to linear)
Loading & Running Gel Generally, followed manufacturer's protocol for running Tris-Glycine 4%-20% mini-gels, Media:LifeTech_NuPAGE_TrisGlycine_Mini_Gels_Cat-EC6025BOX.pdf