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ATF6 Antibody (70B1413.1) - BSA Free

Novus Biologicals, part of Bio-Techne | Catalog # NBP1-40256

Novus Biologicals, part of Bio-Techne

Key Product Details

Validated by

Tagged Protein Expression

Species Reactivity

Human, Mouse, Rat, Porcine, Plant, Rabbit

Applications

Chromatin Immunoprecipitation (ChIP), CyTOF-ready, Flow (Intracellular), Flow Cytometry, Immunoblotting, Immunocytochemistry/Immunofluorescence, Immunohistochemistry, Immunohistochemistry-Frozen, Immunohistochemistry-Paraffin, Immunoprecipitation, Knockout Validated, Western Blot

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG1 kappa Clone # 70B1413.1

Format

BSA Free

Concentration

1.0 mg/ml

Product Specifications

Immunogen

This monoclonal antibody was made against a partial protein containing amino acids 1-273 of human ATF6.

Reactivity Notes

Plant reactivity reported in scientific literature (PMID: 31531232). Use in Plant reported in scientific literature (PMID:31531232).

Specificity

This ATF6 antibody detects both the full length and the cleaved/active protein.

Clonality

Monoclonal

Host

Mouse

Isotype

IgG1 kappa

Description

(1) The ATF6 (IMG-273) has been cited to recognize both full-length and cleaved forms of ATF6. Please refer to the references in the Product Citation list for more comprehensive information.

(2) The ATF6 transfected cell lysate (40210) is recommended as a useful western blot positive control to run in parallel with your experimental samples.

(3) Active/cleaved forms of ATF6 are generated through proteolytic cleavage during ER stress. Different molecular weights have been described for cleaved forms. A sample protocol was first described in Luo and Lee (2002) wherein NIH3T3 cells were treated with the amino acid analogue azetidine (AzC), and full-length and cleaved forms ATF6 were detected with IMG-273. The results show that in addition to the major 90 kDa full length ATF6, protein bands over the range of 50-70 kDa were detectable following AzC treatment (Luo and Lee, 2002: Figure 4A, page 791). Many protocols and other cleaved forms have been described since, please consult the literature for additional information.

(4) For immunofluorescence microscopy, cells were fixed in methanol at -20 degrees C (Thomas et al, 2005). Thomas et al used immunofluorescence to identify ATF6 with IMG-273 in the nucleus.

(5) The active/cleaved 50 kDa nuclear form of ATF6 using IMG-273 has been found to be strongly expressed in certain tumor cell lines derived from B cell lymphoma (DEL), primary effusion lymphoma [BC-3 (ATCC CRL-2277), PEL-SY, HBL-6], lymphoblastic leukemia (DS-1) and multiple myeloma (RPMI-8226, NCI-H929), (Jenner et al. 2003).

(6) Cleaved 60 and 36 kDa ATF6 forms have also been described in the nucleus (Mao et al, 2007).

(7) The ATF6 antibody is reported to be specific for ATF6a, recognizing ATF6a but not ATF6b (Bommiasamy et al, 2009).

(8) In western blots, the binding pattern of ATF6 may vary. Researchers are encouraged to consult the body of literature citing the ATF6 IMG-273 antibody (see Product citation list) for additional information. General ATF6 literature is also helpful. For example, Yoshida (1998) show a multiple band pattern in HeLa in both untreated and stressed cells (Fig 11B). In this early landmark ATF6 publication, multiple bands were seen between the 66 and 116 kDa markers as well as one or more bands between the 45 and 66 kDa markers.

(9) We highly recommend the use of a maximum sensitivity ECL substrate (Femto sensitive) for efficient detection of this antibody in Western blot applications.

Scientific Data Images for ATF6 Antibody (70B1413.1) - BSA Free

Western Blot: ATF6 Antibody (70B1413.1) [NBP1-40256] -

Western Blot: ATF6 Antibody (70B1413.1) [NBP1-40256] - Hypoxia leads to EMT and ER-stress in CRC cells. B. C. Confluent growing SW480 (B) and HCT116 (C) cells were cultured under conditions of normoxia or hypoxia-like conditions (serum free; 100 uM CoCl2, 1-9 h. B-actin (B-act) as loading control. HIF1a was detectable after 3 h of CoCl2 incubation, the amount of the 50 kD-ATF6 fragment, was already enhanced after 1 h of addition of CoCl2. Image collected and cropped by CiteAb from the following publication (//doi.org/10.1371/journal.pone.0087386) licensed under a CC-BY licence.
Western Blot: ATF6 Antibody (70B1413.1) [NBP1-40256] - Analysis of ATF6 in mouse liver tissue using 3 ug/ml of ATF6 antibody and 0.25 ug/ml of GAPDH antibody. Lane A contains 20 ugs of whole mouse liver lysate, lane B contains 20 ugs of total ER fraction, and lane C contains 20 ugs of rough ER fraction. The ATF6 band may represent under glycosylated or cleaved/active ATF6.
Immunocytochemistry/Immunofluorescence: ATF6 Antibody (70B1413.1) [NBP1-40256] - Untreated HeLa cells were fixed in -20C methanol for 10 min, air dried and rehydrated in PBS at room temperature for 5 minutes. Cells were incubated with anti-ATF6 (1:20) for one hour at room temperature. ATF6 reactivity (green) was detected with anti-mouse Dylight-488 secondary antibody. Nuclei were counterstained with DAPI (blue). Note the ER localization of ATF6.

Applications for ATF6 Antibody (70B1413.1) - BSA Free

Application
Recommended Usage

Chromatin Immunoprecipitation (ChIP)

1:10-1:500

Flow Cytometry

1 ug per million cells

Immunocytochemistry/Immunofluorescence

1:10-1:500

Immunohistochemistry

1:10-1:500

Immunohistochemistry-Frozen

1:10-1:500

Immunohistochemistry-Paraffin

1:50

Immunoprecipitation

1:10-1:500

Western Blot

1-5 ug/ml
Application Notes
ICC: See Thomas et al (2005) and Kikuchi et al (2006) for details. Immunohistochemistry (Frozen): See Zhu et al (2008) for details. Immunohistochemistry (Paraffin): See van Kollenburg et al (2006) for details. Immunoprecipitation: See Hong et al, 2004 for details. Use in Immunoblotting reported in scientific literature (PMID 28550308). Knockout validation (PMID: 31531232). In Western blot do not use milk in the diluent as it can inhibit the observed signal. This antibody is CyTOF ready.

Reviewed Applications

Read 3 reviews rated 3.7 using NBP1-40256 in the following applications:

Formulation, Preparation, and Storage

Purification

Protein G purified

Formulation

TBS

Format

BSA Free

Preservative

0.05% Sodium Azide

Concentration

1.0 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: ATF6

ATF6 is a constitutively expressed, endoplasmic reticulum (ER) membrane-anchored transcription factor. ATF6 is a key transcriptional activator of the unfolded protein response (UPR), which allows mammalian cells to maintain cellular homeostasis when they are subjected to environmental and physiological stresses that target the ER (reviewed in Shen, 2005 & Prywes, 2005). The C-terminus of ATF6 is located in the ER lumen and its N-terminal DNA binding domain faces the cytosol. AFT6 plays a key role in the ER stress response by transmitting the ER stress signal across the ER membrane into the nucleus. The induction of new gene expression by ATF6 is an important aspect of the ER stress response. In response to certain stress conditions, ATF6 translocates from the ER to the Golgi. The 90 kDa full-length ATF6 is processed within the Golgi to its active 50 kDa form through sequential cleavage by site-1 and site-2 proteases (S1P and S2P). Proteolytic activation of ATF6 in the ER stress response is a mechanism to regulate membrane-bound factors, and is referred to as regulated intramembrane proteolysis. The N-terminal active ATF6 translocates to the nucleus where it binds to ER stress-response elements in ER stress-response genes (ERSRGs). ATF6 is a potent transcriptional activator of ERSRGs. The fully glycosylated form of ATF6, a 670 amino acid protein, exhibits an electrophoretic mobility of ~90 kDa in denaturing SDS-gels, in part because of the glycosylated modifications. ATF6 has 3 consensus sites for N-linked glycosylation and exists constitutively as a glycosylated protein. Differentially glycosylated ATF6 forms may result from mutations or experimental treatment.

Long Name

Activating Transcription Factor 6

Alternate Names

ACHM7, ATF6 alpha

Entrez Gene IDs

22926 (Human)

Gene Symbol

ATF6

UniProt

Additional ATF6 Products

Product Documents for ATF6 Antibody (70B1413.1) - BSA Free

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot number in the search box below.

Product Specific Notices for ATF6 Antibody (70B1413.1) - BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

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