Human Cell Lines Western Blot

CLS Product Information : Human Cell Lines Western Blot – Kidney carcinoma
Human Cell Lines Western Blot - Kidney carcinoma I
CLS order
number:
309108
Size:
One Blot
Product
description:
Western Blot prepared using CLS lysates of various Kidney Carcinoma cell lines
Source of
proteins
Sample
Triple Color Protein Standard
HEK293 (Human Kidney cell line, adenovirus 5 DNA transformed)
769-P (Human Kidney carcinoma cell line)
ACHN (Human Kidney Adenocarcinoma cell line)
CaKi-1 (Human Kidney clear cell carcinoma cell line)
RCC-FG1 (Human Kidney clear cell carcinoma cell line)
WT-CLS-1 (Human Kidney Wilm’s tumor cell line)
Xxxxx Non-kidney
M
10
1
11
2
3
4
5
6
Lane
M
1
2
3
4
5
6
7
7
M: Triple Color Protein Marker (180kDa, 135kDa,100kDa, 75kDa red,
63kDa, 48 kDa, 35kDa, 25kDa green, 17kDa, 11kDa)
Preparation
Method
Lysates are prepared in 1 x RIPA Lysis Buffer containing protease inhibitors (20mM
Tris-HCl pH 7.5, 150mM NaCl, 1mM Na2EDTA, 1mM EGTA, 1% NP-40, 1%
sodium deoxycholate, 2.5mM sodium pyrophosphate, 1mM ß-glycero-phosphate,
1mM Na3VO4, 1µg/ml leupeptin), 1mM PMSF
Lysates have been run on a 10% precast Tris-Glycine-Polyacrylamide gel,
separation range 30-200kDa
Electrophoresis conditions: 15-25mA, 60min, 1x Laemmli running buffer
Blotting conditions: semi-dry blotting, PVDF membrane, 150mA, 30min,
discontinued buffer sytem (Kyhse-Andersen)
Protein transfer was checked by staining the membrane with reversible protein
staining which results in a turquoise color of the membrane.
Blocking conditions: Non-specific antibody binding is achieved by placing the PVDF
membrane in a 5% bovine serum albumine (BSA) in TBST buffer for 1h at RT.
Applications
Protein detection, antibody staining
Usage
Membrane can be used directly for antibody incubation. We recommend the
following incubation steps: primary antibody of your choice at room temperature for
1-1,5h, 3 x 10min washing steps with TBST buffer, incubation with secondary
antibody of your choice at roomtemperature for 1-1,5h, 3 x 10min washing steps
with TBST buffer.
CLS Cell Lines Service GmbH – Dr.Eckener Strasse 8 – 69214 Eppelheim, Germany - Tel +49(0)6221 700799 – Fax +49(0)6221 700717
[email protected] – www.cell-lines-service.de
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CLS Product Information : Human Cell Lines Western Blot – Kidney carcinoma
For antibody-dilution please follow recommendations on the datasheets of your
antibodys.
Protein detection after the last washing step depends on the conjugation of the
second antibody.
Possible detection reagents are ECL by chemiluminescence for HRP-conjugated
antibodies or BCIP/NBT for alkaline-phosphatase-conjugated antibodies.
Please follow the recommendations on the datasheet of your antibody.
Concentration
30µg of total protein lysate in each lane
Protein
standard
Triple Color Protein Standard 10-180kDa, for details see second page
Storage
The Western Blot membrane should be kept at 4°C until usage. Drying of the
membrane must be avoided.
Important
note
This product is intended for research use only, not for use in human, therapeutic or
diagnostic applications.
CLS Cell Lines Service GmbH – Dr.Eckener Strasse 8 – 69214 Eppelheim, Germany - Tel +49(0)6221 700799 – Fax +49(0)6221 700717
[email protected] – www.cell-lines-service.de
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