Western blot abreview for Anti-Niemann Pick C1 Like 1 antibody [EPR5717]

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Abreviews
Application
Western blot
Sample
Human Cell lysate - whole cell (HEK293T cells over expressing human NPC1L1)
Loading amount
100 µg
Specification
HEK293T cells over expressing human NPC1L1
Gel Running Conditions
Reduced Denaturing (7.5% Tris-Glycine gel)
Blocking step
BSA as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C

Other product details

Dilution
1/1000
Incubation time
24 hour(s) and 0 minute(s) · Temperature: 4°C · Diluent: 5% BSA in TBS-Tween (0.05%)

Secondary antibody

Name
Non-Abcam antibody was used: Goat anti-Rabbit IgG-HRP
Host species: Goat
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Dilution
1/4000

Detection

Detection method
ECL
Exposure
2 second(s)
Bands
Specific: 160 kDa
Positive control
HEK293T cells over expressing human NPC1L1-ACP tagged protein
Negative control
HEK293T cells over expressing mouse NPC1-GFP tagged protein

Additional data

Additional Notes
Human NPC1L1-ACP tagged protein (predicted MW of 160 kDa) and mouse NPC1-GFP tagged protein (predicted MW of 175 kDa) were over expressed in HEK293T cells for 48 hr and lysed in a buffer containing 50mM Tris-HCl pH 7.4, 150 mM NaCl, 5mM MgCl2, 1mM EDTA, 1% Triton X-100, 10% glycerol and protease inhibitors. 100 µg of total cell lysate (with Laemmli buffer) was heated at 50ºC for 25 min. and SDS-PAGE followed by western blotting was performed.

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Submitted Feb 06 2013

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