Primary antibody: BD Pharmingen 555370 (Mouse)
Vendor Link
Source material: Cell line: GBM6
Conditioned media from human glioblastoma patient derived xenograft cell culture GBM6 centrifuged at 10,000 xg for 2hrs and 100,000 xg for 4 hrs. Pellets were resuspended in PBS and protein quantitated using BCA. 10 ug of protein were loaded onto SDS-PAGE.
Secondary antibody: LI-COR 61-6520 (Mouse)
Lysis Conditions: RIPA buffer
at
22°C
for
10 min
Gel Conditions Gel type: Home-made nullselection
Run conditions: 100 V / 0 Amps for 60 min
Reducing conditions: β-mercaptoethanol
Transfer Conditions Membrane type: PVDF
Transfer method: Wet
Transfer buffer: Tris-glycine with 10% methanol
Transfer voltage: 60 V
Transfer current: 0 Amps
Transfer temperature: 4°C
Transfer duration: 4 hr
Blocking Blocking buffer: Intercept Blocking Buffer (LI-COR)
Blocking temperature: 22°C
Duration: 1 hr
Primary Incubation Antibody 1 dilution 1:100
Incubation temperature 22°C
Incubation duration 16 hr
Number of washes 3
Wash buffer TBST
Wash duration 30 min
Secondary Incubation Antibody 2 dilution 1:5,000
Incubation temperature 22°C
Incubation duration 1 hr
Number of washes 3
Wash buffer TBST
Wash duration 30 min
Detection Detection method LI-COR
Expected band size 19 kDa
Result: Weak single band
Background: No background
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Record 3160 author: Alain Charest
Last edit: Tue Mar 19, 2024
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