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AB326976

Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] - BSA and Azide free

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Rabbit Recombinant Monoclonal Cyclin E1 phospho T77 antibody. Carrier free. Suitable for WB, Dot, IP and reacts with Transfected cell lysate - Human, Human, Synthetic peptide - Human samples.

View Alternative Names

CCNE, G1/S-specific cyclin-E1

5 Images
Immunoprecipitation - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] - BSA and Azide free (AB326976)
  • IP

Lab

Immunoprecipitation - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] - BSA and Azide free (AB326976)

This data was developed using ab327426, the same antibody clone in a different buffer formulation.

Cyclin E1 was immunoprecipitated from 0.35 mg HCT 116 (human colorectal carcinoma epithelial cell) treated with 2.5mM Thymidine for 16h whole cell lysate with ab327426 at 1/30 dilution (2μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab327426 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : HCT 116 (human colorectal carcinoma epithelial cell) treated with 2.5mM Thymidine for 16h whole cell lysate
Lane 2 : ab327426 IP in HCT 116 (human colorectal carcinoma epithelial cell) treated with 2.5mM Thymidine for 16h whole cell lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab327426 in HCT 116 treated with 2.5mM Thymidine for 16h whole cell lysate

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Exposure time : 180 seconds.

All lanes:

Immunoprecipitation - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] (ab327426) at 1/1000 dilution

Lane 1:

HCT 116 (human colorectal carcinoma epithelial cell) treated with 2.5mM Thymidine for 16h whole cell lysate at 10 µg

Lane 2:

ab327426 at 1/30 IP in HCT 116 (human colorectal carcinoma epithelial cell) treated with 2.5mM Thymidine for 16h whole cell lysate

Lane 3:

Rabbit monoclonal IgG (<a href='/en-us/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab327426 in HCT 116 treated with 2.5mM Thymidine for 16h whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 35-50 kDa

false

Exposure time: 180s

Western blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] - BSA and Azide free (AB326976)
  • WB

Lab

Western blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] - BSA and Azide free (AB326976)

This data was developed using ab327426, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) (1 : 200000) (36KDa).

All lanes:

Western blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] (ab327426) at 1/1000 dilution

Lane 1:

Untreated A549 (human lung carcinoma epithelial cell) whole cell lysate (untreated membrane) at 50 µg

Lane 2:

A549 treated with 2.5mM Thymidine for 16h whole cell lysate (untreated membrane) at 50 µg

Lane 3:

A549 treated with 2.5mM Thymidine for 16h whole cell lysate (alkaline phosphatase treated membrane) at 50 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 35-50 kDa,36 kDa

false

Exposure time: 48s

Western blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] - BSA and Azide free (AB326976)
  • WB

Lab

Western blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] - BSA and Azide free (AB326976)

This data was developed using ab327426, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

In Western blot, ab327426 was shown to bind specifically to CCNE1. Target of interest was observed at 35-50 kD in wild-type HCT116 cell lysates (lanes 1-2) with no signal observed at this size in the CCNE1 knockout cell line (lanes 3-4) (lanes 3-4, knockout cell line ab286494).

The identity of the higher MW band at approximately 150 kDa is unknown.

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) (1 : 10000) (124KDa).

All lanes:

Western blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] (ab327426) at 1/1000 dilution

Lane 1:

Untreated HCT 116 (human colorectal carcinoma epithelial cell) whole cell lysate (untreated membrane) at 50 µg

Lane 2:

HCT 116 treated with 2.5mM Thymidine for 16h whole cell lysate (untreated membrane) at 50 µg

Lane 3:

CCNE1 knockout HCT116 whole cell lysate (untreated membrane) at 50 µg

Lane 4:

CCNE1 knockout HCT116 treated with 2.5mM Thymidine for 16h) whole cell lysate (untreated membrane) at 50 µg

Lane 5:

Untreated HCT 116 whole cell lysate (alkaline phosphatase treated membrane) at 50 µg

Lane 6:

HCT 116 treated with 2.5mM Thymidine for 16h whole cell lysate (alkaline phosphatase treated membrane) at 50 µg

Lane 7:

CCNE1 knockout HCT116 whole cell lysate (alkaline phosphatase treated membrane) at 50 µg

Lane 8:

CCNE1 knockout HCT116 treated with 2.5mM Thymidine for 16h whole cell lysate (alkaline phosphatase treated membrane) at 50 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 35-50 kDa,124 kDa

false

Exposure time: 180s

Western blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] - BSA and Azide free (AB326976)
  • WB

Lab

Western blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] - BSA and Azide free (AB326976)

This data was developed using ab327426, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) (1 : 200000) (36KDa) ;Anti-6X His tag antibody [EPR20547]-CHIP Grade (ab213204) (1 : 5000)(35-50KDa).

All lanes:

Western blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] (ab327426) at 1/5000 dilution

Lane 1:

293T cells transfected with a human wild-type CCNE1 expression vector containing a His-tag, whole cell lysate at 20 µg

Lane 2:

293T cells transfected with a human CCNE1 (T77A) expression vector containing a His-tag, whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/200000 dilution

Observed band size: 35-50 kDa,36 kDa

false

Exposure time: 15s

Dot Blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] - BSA and Azide free (AB326976)
  • Dot

Lab

Dot Blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] - BSA and Azide free (AB326976)

This data was developed using ab327426, the same antibody clone in a different buffer formulation.

Dot blot analysis of Cyclin E1 using ab327426 at 1 : 1000 (0.504 μg/ml) followed by a Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1 : 100,000 dilution.

Lane 1 : CCNE1 (phospho T77) peptide

Lane 2 : CCNE1 (phospho T77) peptide

Lane 3 : CCNE1 non-phospho peptide

Lane 4 : CCNE1 (phospho S73+ T77) peptide

Lane 5 : CCNE1 (phospho S73) peptide

Exposure time : 180 seconds.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

All lanes:

Dot Blot - Anti-Cyclin E1(Phospho T77) antibody [EPR28889-10] (ab327426) at 1/1000 dilution

Lanes 1 - 2:

CCNE1 (phospho T77) peptide

Lane 3:

CCNE1 non-phospho peptide

Lane 4:

CCNE1 (phospho S73+ T77) peptide

Lane 5:

CCNE1 (phospho S73) peptide

Secondary

All lanes:

Dot Blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

false

Exposure time: 180s

Key facts

Host species

Rabbit

Clonality

Monoclonal

Clone number

EPR28889-10

Isotype

IgG

Carrier free

Yes

Reacts with

Human

Applications

Dot, WB, IP

applications

Immunogen

The exact immunogen used to generate this antibody is proprietary information.

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Reactivity data

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Product details

ab326976 is the carrier-free version of ab327426

Patented technology
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

What are the advantages of a recombinant monoclonal antibody?
This product is a recombinant monoclonal antibody, which offers several advantages including:

  • - High batch-to-batch consistency and reproducibility
  • - Improved sensitivity and specificity
  • - Long-term security of supply
  • - Animal-free batch production

For more information, read more on recombinant antibodies.

Conjugation ready
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.

Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with 1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

Compatibility
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.

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Properties and storage information

Form
Liquid
Purification technique
Affinity purification Protein A
Storage buffer
pH: 7.2 - 7.4 Constituents: PBS
Shipped at conditions
Blue Ice
Appropriate short-term storage conditions
+4°C
Appropriate long-term storage conditions
+4°C
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Product protocols

For this product, it's our understanding that no specific protocols are required. You can visit:
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Target data

Essential for the control of the cell cycle at the G1/S (start) transition.
See full target information CCNE1 phospho T77
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Product promise

We are committed to supporting your work with high-quality reagents, and we're here for you every step of the way. In the unlikely event that one of our products does not perform as expected, you're protected by our Product Promise.
For full details, please see our Terms & Conditions

Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.

For licensing inquiries, please contact partnerships@abcam.com