EV Antibody Database

Western Blot Antibody

Target protein:

CD81

Primary antibody:

Santa Cruz Biotech 23962 (Human)

No notes

Source material:

Cell line: DiFi

DiFi conditioned media was produced by Dr. Jeffrey Franklin of Vanderbilt University. Media was fractionated via Density-Gradient Ultracentrifugation, resulting in 12x 1mL fractions. Fractions 5-12 were run on a WB to detect CD81 activity.

Secondary antibody:

Santa Cruz Biotech sc-516102 (Mouse)

No notes

Lysis Conditions:

Laemmli Buffer
at
95°C
for
5 min

Gel Conditions

Minigel, 4 % acrylamide

 

Gel type:

BioRad 4568094

Run conditions:

120 V / Variable Amps for 50min

Protein loading:

10 𝜇g

Reducing conditions:

Non-reducing

No notes

Transfer Conditions

Membrane type:

PVDF

Transfer method:

Wet

Transfer buffer:

Tris-glycine with 10% methanol

Transfer voltage:

Variable V

Transfer current:

.35 Amps

Transfer temperature:

4°C

Transfer duration:

2 hr

Blocking

Blocking buffer:

5% milk, 0% BSA, No serum

Blocking temperature:

22°C

Duration:

1 hr

Primary Incubation

Antibody 1 dilution

1:500

Incubation temperature

4°C

Incubation duration

16 hr

Number of washes

4

Wash buffer

TBST, 0.1% Tween-20

Wash duration

5 min

Secondary Incubation

Antibody 2 dilution

1:1,000

Incubation temperature

22°C

Incubation duration

1 hr

Number of washes

4

Wash buffer

TBST, 0.1% Tween-20

Wash duration

5 min

Detection

Detection method

Enhanced chemiluminescence (ECL)

Expected band size

25 kDa

Result:

Strong single band

Background:

No background

Related Publications

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No comments

Record 9806 author: Martin Ng

Author Information

Email:martin.ng@ncire.org
Lab Head:
Institution:
Website:https://raffailab.ucsf.edu/
Last edit: Thu Mar 14, 2024