Recombinant Anti-5T4 antibody [EPR5530] KO Tested (ab129058)

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Western blot - Anti-5T4 antibody [EPR5530] (ab129058)

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [EPR5530] to 5T4
Suitable for: WB, IP
Knockout validated
Reacts with: Rat, Human

Carrier Free HRP

Product name

Anti-5T4 antibody [EPR5530]
See all 5T4 primary antibodies
Description

Rabbit monoclonal [EPR5530] to 5T4
Host species

Rabbit
Tested applications

Suitable for: WB, IPmore details
Unsuitable for: Flow Cyt,ICC/IF or IHC-P
Species reactivity

Reacts with: Rat, Human
Predicted to work with: Mouse
Immunogen

Synthetic peptide within Human 5T4 aa 350-450 (intracellular). The exact sequence is proprietary.
Positive control
- HEK-293T, HT 1376, HeLa, JAR and MCF7 cell lysates; MCF7 cells.
General notes
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 production
For more information see here.
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.
We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

Form

Liquid
Storage instructions

Shipped at 4°C. Store at -20°C. Stable for 12 months at -20°C.
Storage buffer

pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR5530
Isotype

IgG
Research areas

Alternative Versions
- Anti-5T4 antibody [EPR5530] - BSA and Azide free (ab189508)
- HRP Anti-5T4 antibody [EPR5530] (ab199547)
Isotype control
- Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
KO cell lines
- Human TPBG knockout MCF7 cell line (ab269499)
KO cell lysates
- Human TPBG knockout MCF7 cell lysate (ab269661)
Positive Controls
- MCF7 membrane extract lysate (ab29539)
- HeLa membrane extract lysate (ab29547)

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab129058 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application	Abreviews	Notes
WB		1/1000 - 1/10000. Detects a band of approximately 72-80 kDa (predicted molecular weight: 46 kDa).
IP		1/80.

Notes
WB 1/1000 - 1/10000. Detects a band of approximately 72-80 kDa (predicted molecular weight: 46 kDa).
IP 1/80.

Application notes

Is unsuitable for Flow Cyt,ICC/IF or IHC-P.

Tissue specificity

Expressed by all types of trophoblasts as early as 9 weeks of development. Specific for trophoblastic cells except for amniotic epithelium. In adult tissues, the expression is limited to a few epithelial cell types but is found on a variety of carcinoma.
Sequence similarities

Contains 6 LRR (leucine-rich) repeats.
Contains 1 LRRCT domain.
Contains 1 LRRNT domain.
Cellular localization

Membrane.
Target information above from: UniProt accession Q13641 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
Database links
- Entrez Gene: 7162 Human
- Entrez Gene: 21983 Mouse
- Entrez Gene: 83684 Rat
- Omim: 190920 Human
- SwissProt: Q13641 Human
- SwissProt: Q9Z0L0 Mouse
- SwissProt: Q5PQV5 Rat
- Unigene: 82128 Human
- Unigene: 20864 Mouse
- Unigene: 484180 Mouse
- Unigene: 43122 Rat
see all
Form

5T4 is a 72 kDa transmembrane glycoprotein that exhibits restricted expression in human and mouse adult tissues, is upregulated on many carcinomas and tumour expression correlates with poorer clinical outcome in ovarian, gastric and colorectal cancers. mES cells lack cell surface expression of the 5T4 antigen, and both protein and mRNA are rapidly upregulated following induction of differentiation. This proteins expression correlates with downregulation of OCT-4 and Tra-1–60 (Ward et al., 2006).
Alternative names
- 5T4 antibody
- 5T4 oncofetal antigen antibody
- 5T4 oncofetal trophoblast glycoprotein antibody
- 5T4 oncotrophoblast glycoprotein antibody
- 5T4AG antibody
- AW495680 antibody
- M6P1 antibody
- TPBG antibody
- TPBG_HUMAN antibody
- Trophoblast glycoprotein antibody
see all

Western blot - Anti-5T4 antibody [EPR5530] (ab129058)

All lanes : Anti-5T4 antibody [EPR5530] (ab129058) at 1/1000 dilution

Lane 1 : Wild-type MCF7 (Human breast adenocarcinoma cell line) whole cell lysate
Lane 2 : TPBG knockout MCF7 (Human breast adenocarcinoma cell line) whole cell lysate
Lane 3 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 4 : HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 46 kDa
Observed band size: 85 kDa why is the actual band size different from the predicted?

Lanes 1 - 4: Merged signal (red and green). Green - ab129058 observed at 85 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.

ab129058 was shown to react with 5T4 in wild-type MCF7 cells in Western blot with loss of signal observed in TPBG knockout cell line ab269499 (knockout cell lysate ab269661). Wild-type MCF7 and TPBG knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in before incubation with ab129058 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
Western blot - Anti-5T4 antibody [EPR5530] (ab129058)

Anti-5T4 antibody [EPR5530] (ab129058) at 1/5000 dilution (purified) + Rat brain at 20 µg

Secondary
HRP goat-anti rabbit (H+L) at 1/1000 dilution

Predicted band size: 46 kDa

Blocking buffer: 5% NFDM/TBST
Dilution buffer: 5% NFDM/TBST
Western blot - Anti-5T4 antibody [EPR5530] (ab129058)

Anti-5T4 antibody [EPR5530] (ab129058) at 1/2000 dilution (unpurified) + Rat brain at 20 µg

Secondary
HRP goat anti-rabbit (H+L) at 1/1000 dilution

Predicted band size: 46 kDa

Blocking buffer: 5% NFDM/TBST
Dilution buffer: 5% NFDM/TBST
Western blot - Anti-5T4 antibody [EPR5530] (ab129058)

All lanes : Anti-5T4 antibody [EPR5530] (ab129058) at 1/5000 dilution (purified)

Lane 1 : HT-1376 cell lysate
Lane 2 : HeLa cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : HRP goat anti-rabbit (H+L) at 1/1000 dilution

Predicted band size: 46 kDa

Blocking buffer: 5% NFDM/TBST
Dilution buffer: 5% NFDM/TBST
Western blot - Anti-5T4 antibody [EPR5530] (ab129058)

All lanes : Anti-5T4 antibody [EPR5530] (ab129058) at 1/2000 dilution (unpurified)

Lane 1 : HT-1376 cell lysate
Lane 2 : HeLa cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : HRP goat anti-rabbit (H+L) at 1/1000 dilution

Predicted band size: 46 kDa

Blocking buffer: 5% NFDM/TBST
Dilution buffer: 5% NFDM/TBST
Western blot - Anti-5T4 antibody [EPR5530] (ab129058)

All lanes : Anti-5T4 antibody [EPR5530] (ab129058) at 1/1000 dilution (unpurified)

Lane 1 : HT 1376 cell lysate
Lane 2 : HeLa cell lysate
Lane 3 : JAR cell lysate
Lane 4 : MCF7 cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat anti-rabbit HRP at 1/2000 dilution

Predicted band size: 46 kDa
Immunoprecipitation - Anti-5T4 antibody [EPR5530] (ab129058)

ab129058 (unpurified) at 1/50 immunoprecipitating 5T4 in MCF-7 (Lane 1). For western blotting, a HRP goat anti-rabbit was used as the secondary antibody (1/1000).
Blocking buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM /TBST.
Immunoprecipitation - Anti-5T4 antibody [EPR5530] (ab129058)

ab129058 (unpurified) at 1/50 immunoprecipitating 5T4 in MCF-7 (Lane 1). For western blotting, a HRP goat anti-rabbit was used as the secondary antibody (1/1000).
Blocking buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM /TBST.
Anti-5T4 antibody [EPR5530] (ab129058)

Western blot protocols
Immunoprecipitation protocols

Click here to view the general protocols

Datasheet download

Download

Publishing research using ab129058? Please let us know so that we can cite the reference in this datasheet.

ab129058 has been referenced in 4 publications.

Wakeham CM et al. Expression and distribution of trophoblast glycoprotein in the mouse retina. J Comp Neurol 528:1660-1671 (2020). PubMed: 31891182
Wakeham CM et al. Identification of PKCa-dependent phosphoproteins in mouse retina. J Proteomics 206:103423 (2019). PubMed: 31255707
Puchert M et al. The 5T4 oncofetal glycoprotein does not act as a general organizer of the CXCL12 system in cancer cells. Exp Cell Res 364:175-183 (2018). PubMed: 29408206
Shor B et al. Enhanced Antitumor Activity of an Anti-5T4 Antibody-Drug Conjugate in Combination with PI3K/mTOR inhibitors or Taxanes. Clin Cancer Res 22:383-94 (2016). WB ; Human . PubMed: 26319086

Customer reviews and Q&As

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Application

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)

Sample

Human Tissue sections (placenta)

Antigen retrieval step

Heat mediated - Buffer/Enzyme Used: pH 9.0

Permeabilization

Specification

placenta

Blocking step

Serum as blocking agent for 15 minute(s) · Concentration: 5% · Temperature: 20°C

Fixative

Formaldehyde

The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

DR. Lennart Olsson

Verified customer

Submitted Apr 25 2019

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
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Key features and details

Related conjugates and formulations

Overview

Product name

Description

Host species

Tested applications

Species reactivity

Immunogen

Positive control

General notes

Properties

Form

Storage instructions

Storage buffer

Purity

Clonality

Clone number

Isotype

Research areas

Associated products

Alternative Versions

Isotype control

KO cell lines

KO cell lysates

Positive Controls

Applications

The Abpromise guarantee

Target

Tissue specificity

Sequence similarities

Cellular localization

Database links

Form

Alternative names

Images

Protocols

Datasheets and documents

Datasheet download

References (4)

Customer reviews and Q&As

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-5T4 antibody [EPR5530]