Western Blot Antibody Customer Review for p63 Polyclonal Antibody (STJ94912)

HOW DID OUR ANTIBODY PERFORM?

ANTIBODY CUSTOMER REVIEW: p63 Polyclonal Antibody (STJ94912)

What was used?Primary antibody: STJ94912 p63 antibody

Provider: St John's Laboratory Ltd.

Dilution ratio: 1:500

Application: Western Blot

Materials for validation: Human corneal limbal stem cells

Click for product data sheet PDF

What Was The Protocol?Treatment of Materials When cells were cultured to 70%-80% plating density, lysis buffer with protease inhibitors

(Roche) and equal volume of 2X loading buffer were added for 10 minutes in a water bath at 100˚C.

Gel Electrophoresis 10% polyacrylamide gel, voltage 80V for 15 minutes, then 140 V for 45 minutes (total 60 minutes).

Transfer 0.45 µm NC membrane, 1 piece of gel (wet transfer), constant voltage 100 V 60 minutes.

Blocking Odyssey Blocking Buffer (PBS) for 1 hour.

Primary Antibody Probing Primary antibody was added to the solution of Odyssey Blocking Buffer (PBS) plus 0.01% Tween 20, 4˚C overnight.

Membrane Wash Odyssey Blocking Buffer (PBS) plus 0.01% Tween-20 for 4 times.

Secondary Antibody Probing Secondary antibody was added to the solution of Odyssey Blocking Buffer (PBS) plus 0.01% Tween-20 and 0.02% SDS with 1:10000 dilution ratio, for 1 hour.

Membrane Wash Odyssey Blocking Buffer (PBS) plus 0.01% Tween-20 for 4 times.

Visualization Odyssey CLx Near-Infrared Imaging System.

What was the protocol?10% polyacrylamide gel, voltage 80V for 15 minutes, then 140 V for 45 minutes (total 60 minutes).

Transfer information: 0.45 µm NC membrane, 1 piece of gel (wet transfer), constant voltage 100 V 60 minutes.

No. Antigen Loading amount

Primary antibody

Primary antibody

dilution ratio

Secondaryantibody

dilution ratioTarget band KD Visualization

time

1P0 human

corneal limbal stem cells (P0)

20µg (total protein)

STJ94912 p63 antibody

1:500

1:0000 ~77KD 60s

2P1 human

corneal limbal stem cells (P1)

20µg (total protein) ~77KD 60s1:0000

What were the results?

Figure: Lane1: P0; Lane 2: P1

What did the customer think?

Antibody Specificity:

Antibody RPPA Rating:

Testing results were provided independently by F. Abidin at Newcastle University. They stated “In human corneal limbal stem cells, there were target bands detected by STJ94912 p63 antibody”

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