ATF6 Antibody (70B1413.1) - BSA Free

Novus Biologicals | Catalog # NBP1-40256

Novus Biologicals

Key Product Details

Validated by

Biological Validation

Species Reactivity

Validated:

Human, Mouse, Rat, Porcine, Plant, Rabbit

Cited:

Human, Mouse, Rat, Porcine, Plant, Rabbit

Applications

Validated:

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

Cited:

Knockout Validated, Immunohistochemistry, Immunohistochemistry-Paraffin, Immunohistochemistry-Frozen, Western Blot, Immunoblotting, Immunocytochemistry/ Immunofluorescence, Simple Western, Chemotaxis, IF/IHC

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG1 kappa Clone # 70B1413.1

Format

BSA Free
View all ATF6 Primary Antibodies »

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] -

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

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

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) - BSA Free [NBP1-40256]

Immunocytochemistry/ Immunofluorescence: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256]

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.
Immunohistochemistry-Paraffin: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256]

Immunohistochemistry-Paraffin: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256]

Immunohistochemistry-Paraffin: ATF6 Antibody (70B1413.1) [NBP1-40256] - Analysis of ATF6 antibody on Human placenta. Fixed paraffin-embedded sections. Antibody dilution 1:50. Incubated overnight in 4C. Image from verified customer review.
Flow (Intracellular): ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256]

Flow (Intracellular): ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256]

Flow (Intracellular): ATF6 Antibody (70B1413.1) [NBP1-40256] - An intracellular stain was performed on HeLa cells with ATF6 Antibody (70B1413.1) NBP1-40256F (blue) and a matched isotype control (orange). Cells were fixed with 4% PFA and then permeablized with 0.1% saponin. Cells were incubated in an antibody dilution of 10 ug/mL for 30 minutes at room temperature. Both antibodies were conjugated to FITC.. Using the FITC format of this antibody.
Western Blot: ATF6 Antibody (70B1413.1)BSA Free [NBP1-40256]

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

Western Blot: ATF6 Antibody (70B1413.1) [NBP1-40256] - Lane 1: 293 cells transfected with full-length ATF6.
Lane 2: 293 cells transfected with partial length ATF6 (amino acids 1-373).
Lane 3: Untransfected 293 cells.

Western blots were probed with 4 ug/ml of the ATF6 monoclonal antibody and visualized with PicoTect Western Blot Chemiluminescence Substrate (10087K). Film was exposed for 1 min. The top arrow corresponds to the 90 kDa form of ATF6 described as full-length in the literature. The human full-length and partial length ATF6 plasmids are described in Luo and Lee (2002).
Immunohistochemistry-Paraffin: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256]

Immunohistochemistry-Paraffin: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256]

Immunohistochemistry-Paraffin: ATF6 Antibody (70B1413.1) [NBP1-40256] - Human placenta, followed by biotinylated horse anti-mouse IgG secondary antibody, alkaline phosphatase-streptavidin and chromogen. Dilution 10ug/ml
Flow Cytometry: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256]

Flow Cytometry: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256]

Flow Cytometry: ATF6 Antibody (70B1413.1) [NBP1-40256] - Intracellular flow cytometric staining of 1 x 10^6 MCF-7 cells using ATF6 antibody (dark blue). Isotype control shown in orange. An antibody concentration of 1 ug/1x10^6 cells was used.
ATF6 Antibody (70B1413.1) - BSA Free

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

Western Blot: ATF6 Antibody (70B1413.1) [NBP1-40256] - Lysate of DU145 cells. Image from verified customer review.
ATF6 Antibody (70B1413.1) - BSA Free

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

Western Blot: ATF6 Antibody (70B1413.1) [NBP1-40256] - Lysate of PC-3 cells. Image from verified customer review.
ATF6 Antibody (70B1413.1) - BSA Free

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

Expression of ER stress-associated transcription factors in NASH, NAFL and normal liver tissues beta -Actin was used as an internal control. Horizontal lines represent means of densitometry signals from the western blot analyses for all tissue groups. #, significant differences in signals between NASH and normal liver tissues (P < 0.05). Data for NAFL tissues were not used for statistical comparisons because of limited sample number (n = 2).
ATF6 Antibody (70B1413.1) - BSA Free

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

Abnormal Paneth cells show ER stress.(A, B) Representative transmission electron microscopy images of Paneth cells at the base of ileal crypts in (A) ICR & (B) SAMP1/YitFc mice. Scale bars indicate 2 μm. (C, D) Quantitative analysis of (C) granule number & (D) ER lumen diameter in Paneth cells (n = 3/each week for SAMP1/YitFc mice). For the measurements, three Paneth cells were randomly selected from each mouse. (E) SDS–PAGE Western blot analysis of ER stress markers, pIRE1 alpha, ATF4, cleaved-ATF6, & GRP78 in ileal crypts (n = 4/each group). Total-IRE1 alpha & HPRT1 was used as loading control. (F) Relative expression level of ER stress markers calculated from the band intensity. Error bars represent mean ± SEM. (C, D, F) Statistical significance was evaluated by t test in (C, D), & one-way ANOVA followed by Tukey’s post hoc test in (F). P < 0.05 was considered statistically significant. *P < 0.05, †P < 0.01, §P < 0.001. E, ER; G, granules; N, nucleus; n.s., not significant. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32345659), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

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

Western Blot: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - The ISR is active in the retinas of rd16 mice.Western blots & corresponding graphs of ISR markers in the retinas of rd16 mice at P15 (a) & P20 (b). At P15, p-eIF2 alpha, ATF4, CHOP, & TRB3 are significantly elevated. At P20, levels of p-eIF2 alpha, ATF4, CHOP, & GADD34 are increased. Markers of the UPR are elevated in rd16 retinas. BiP & cleaved (~50 kD) ATF6 are upregulated in the retinas of rd16 mice at P15 (c) & P20 (d). Relative density measurements correspond to the intensities of the immunoblotting bands or lanes normalized to an internal control. Data are shown as mean ± SEM. a.u. arbitrary units. P15 n = 3. P20 n = 4. *p < 0.05, **p < 0.01, ***p < 0.001 Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/29706649), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

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

Western Blot: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - Expression of ER stress-associated transcription factors in NASH, NAFL & normal liver tissues beta -Actin was used as an internal control. Horizontal lines represent means of densitometry signals from the western blot analyses for all tissue groups. #, significant differences in signals between NASH & normal liver tissues (P < 0.05). Data for NAFL tissues were not used for statistical comparisons because of limited sample number (n = 2). Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/28968997), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

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

Western Blot: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - Restoration of Plk3 induction rescues ATF2 phosphorylation & ER stress resolution in Lrh-1LKO mice, & loss or gain of ATF2 transcriptional activity also alters ER stress resolution capacity.(A) Primary hepatocytes prepared from Lrh-1f/fand Lrh-1LKO mice & transduced with Ad-Plk3 or Ad-control at a MOI of 100. Cells treated 36 hr later with vehicle or tunicamycin (TM) (0.01 µg/ml) & doxycycline (1 µg/ml) to induce Plk3 or LacZ control. Nuclear protein was obtained at timepoints indicated & immunoblotted for UPR transcription factors, with TBP as a loading control. Results representative of 3 independent experiments. Image collected & cropped by CiteAb from the following publication (https://elifesciences.org/articles/01694), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

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

Western Blot: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - The ISR is active in the retinas of rd16 mice.Western blots & corresponding graphs of ISR markers in the retinas of rd16 mice at P15 (a) & P20 (b). At P15, p-eIF2 alpha, ATF4, CHOP, & TRB3 are significantly elevated. At P20, levels of p-eIF2 alpha, ATF4, CHOP, & GADD34 are increased. Markers of the UPR are elevated in rd16 retinas. BiP & cleaved (~50 kD) ATF6 are upregulated in the retinas of rd16 mice at P15 (c) & P20 (d). Relative density measurements correspond to the intensities of the immunoblotting bands or lanes normalized to an internal control. Data are shown as mean ± SEM. a.u. arbitrary units. P15 n = 3. P20 n = 4. *p < 0.05, **p < 0.01, ***p < 0.001 Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/29706649), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

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

Western Blot: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - APPswe induction of OB-senescence via ER stress.a Heat map of differentially expressed ER stress or anti-stress related genes identified by RNA-seq in control (OCN-Cre; Ai9) & TgAPPsweOCN; Ai9 Td+ OB-progenitors (detail analysis was described in Methods). b RT-PCR analysis of ER stress-related genes Grp78, Atf6, Hsp90b1, Eif2ak3, Ern1, Hsp90aa1, & Hspa2 & anti-stress related gene Sirt3 gene expression in purified Td+ BMSCs from 6-MO control (OCN-Cre; Ai9) & TgAPPsweOCN; Ai9 mice, *p < 0.05, **p < 0.01, ***p < 0.001, mean ± SD, n = 3, Mann–Whitney U test. c Western blot analysis of indicated protein expression in BMSCs from mice with indicated genotypes (at 6-MO). GAPDH was used as a loading control. d Quantification of data in c, *p < 0.05, **p < 0.01. mean ± SD, n = 4, Student’s t test. e Western blot analysis of indicated protein expression in BMSCs from 6-MO control & TgAPPsweOCN with or without 0.25 mM 4-PBA (4-Phenylbutyric acid) treatment. f Quantification analyses of the data in e, *p < 0.05, n = 3. g SA-beta -gal staining of 6-MO control & TgAPPsweOCN BMSCs with vehicle (Veh)(PBS) & 4-PBA treatment, respectively, scale bar, 20 µm. h Quantification of SA-beta -gal+ cell densities in g (mean ± SD; n = 5, **p < 0.01, ***p < 0.001). Two-way analysis of variance test was used in f & h. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/34824365), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

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

Western Blot: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - Disturbance of ER homeostasis in ADTKD-UMOD.(A) ADTKD-UMOD is characterized by maturation & trafficking defect of mutant UMOD & intracellular accumulation of UMOD in TAL cells. UMOD immunolocalization revealed a diffuse cytoplasmic staining with enforcement of the luminal membrane in TAL cells of a wild-type mouse. In contrast, TAL cells of an UmodC93F mutant mouse displayed a strong paranuclear immunopositivity for UMOD. Wild-type: Umodwt mouse; UmodC93F: homozygous UmodC93F mutant mouse. Age of mice analysed: four months. Chromogen: DAB, nuclear staining: haemalum. (B) Heat map of relative expression values (z scores) showed differential abundance of several proteins localized in the ER. (C) In the outer medulla of Umod mutant mice of both mouse lines, a strong accumulation of immature UMOD was present. (D) Protein abundances of BiP, phospho-eIF2 alpha, eIF2 alpha, ATF4, both full-length (§) & cleaved activated (#) ATF6, & CHOP were increased in Umod mutant mice compared to wild-type mice. Signal intensities were corrected for GAPDH signal intensities of the same PVDF-membrane, which was stripped several times to facilitate the detection of multiple proteins. Mean of protein abundance of wild-type mice was set on a value of 1 [mean (wild-type) = 1]. Data are shown as means ± SD. One-way ANOVA with Newman-Keuls’s post hoc test: p vs. wild-type, *p < 0.05; **p < 0.01; ***p < 0.001. Image collected & cropped by CiteAb from the following publication (https://www.nature.com/articles/srep42970), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

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

Western Blot: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - The effect of Ezetimibe on unfolded protein response (UPR) gene expression in THP-1 cells exposed to ischemia-reperfusion (IR). (a) The mRNA expression of activating transcription factor 6 (ATF6) & CCAAT-enhancer-binding protein homologous protein (CHOP). (b) Representative Western blot analyses for the indicated proteins. (c,d) The average quantification of ATF6 & CHOP obtained by the densitometric analysis of three independent experiments. mRNA was analyzed by quantitative real-time PCR; normalized gene expression levels are given as the ratio between the mean value for the target gene & that for beta -actin in each sample. Data are expressed as mean ± SD. * p < 0.01 vs. control (up-regulation); ** p < 0.01 vs. IR; §p < 0.01 vs. control (down-regulation). Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32340270), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

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

Western Blot: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - APPswe induction of OB-senescence via ER stress.a Heat map of differentially expressed ER stress or anti-stress related genes identified by RNA-seq in control (OCN-Cre; Ai9) & TgAPPsweOCN; Ai9 Td+ OB-progenitors (detail analysis was described in Methods). b RT-PCR analysis of ER stress-related genes Grp78, Atf6, Hsp90b1, Eif2ak3, Ern1, Hsp90aa1, & Hspa2 & anti-stress related gene Sirt3 gene expression in purified Td+ BMSCs from 6-MO control (OCN-Cre; Ai9) & TgAPPsweOCN; Ai9 mice, *p < 0.05, **p < 0.01, ***p < 0.001, mean ± SD, n = 3, Mann–Whitney U test. c Western blot analysis of indicated protein expression in BMSCs from mice with indicated genotypes (at 6-MO). GAPDH was used as a loading control. d Quantification of data in c, *p < 0.05, **p < 0.01. mean ± SD, n = 4, Student’s t test. e Western blot analysis of indicated protein expression in BMSCs from 6-MO control & TgAPPsweOCN with or without 0.25 mM 4-PBA (4-Phenylbutyric acid) treatment. f Quantification analyses of the data in e, *p < 0.05, n = 3. g SA-beta -gal staining of 6-MO control & TgAPPsweOCN BMSCs with vehicle (Veh)(PBS) & 4-PBA treatment, respectively, scale bar, 20 µm. h Quantification of SA-beta -gal+ cell densities in g (mean ± SD; n = 5, **p < 0.01, ***p < 0.001). Two-way analysis of variance test was used in f & h. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/34824365), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

Immunocytochemistry/ Immunofluorescence: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] -

Immunocytochemistry/ Immunofluorescence: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - Effect of rapamycin on markers of the UPR pathways(A) Cells were incubated in medium alone (0.05% FBS), with 3 nM IGF-1 or with both 3 nM IGF-1 & 10 nM rapamycin (IGF-1 + rapamycin). Total protein extracts prepared from cells incubated for 24 h in those conditions were subjected to Western Blot analysis. Protein expression levels were assessed for phosphorylated or total forms of PERK, eIF2 alpha & for CHOP, BiP, XBP1-s & XBP1-u proteins. Efficiency of rapamycin to inhibit mTORC1 pathway was also checked by immunoblot with phosphorylated & total forms of p70S6K1 & 4E-BP1. alpha -tubulin was used as internal control. (B & C) Cells were incubated with 10nM rapamycin for 1 h (B) or 24 h (C). Immunoblots for phosphorylated & total forms of PERK, eIF2 alpha, p70S6K1 & 4E-BP1 or for ATF4, CHOP, BiP, XBP1-s & XBP1-u protein expression were performed. alpha -tubulin was used as internal control. Blots of P-p70S6K1, p70S6K1, P-PERK, PERK, & alpha -tubulin of Figure 4B & blots of Figure 4C have been performed on the same electrophoresis gel, but cut & reconstituted. (D) Cells were incubated in medium alone (Ctrl) or with 10 nM rapamycin for 24 h. Nuclear localization of ATF6 was assessed using immunofluorescence with ATF6-antibody (red) & Hoechst dye. Magnification ×1000. (E) Bar graphs were obtained by quantification of ATF6 nuclear staining. Results are representative of at least 3 experiments. Image collected & cropped by CiteAb from the following publication (https://www.oncotarget.com/lookup/doi/10.18632/oncotarget.15469), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

Immunocytochemistry/ Immunofluorescence: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] -

Immunocytochemistry/ Immunofluorescence: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - Activation of the UPR during hypoxia or glucose depletionSTC-1 & GluTag cells were subjected to hypoxia (1%) or cultivated with decreasing concentration of glucose i.e. 25, 5 or 1 mM, for 24 h. (A) & (C) Protein expression level of phosphorylated & total forms of PERK, eIF2 alpha & ATF4, CHOP, BiP & C-Caspase 3 protein expression was examined using immunoblots, during hypoxia (A) & glucose depletion (C). alpha -tubulin was used as internal control. Blots of Figure 2A have been performed on the same electrophoresis gel, but cut & reconstituted. (B) & (D) XBP1 mRNA splicing was analyzed by RT-PCR after Pst1 digestion: XBP1-u, unspliced; XBP1-h, hybrid; XBP1-s, spliced variant of XBP1; *, XBP1-u mRNA fragments after Pst1 digestion. (E) ATF6 nuclear localization was assessed in STC-1 cells using immunofluorescence with anti-ATF6 antibody & hoechst dye. Magnification x1000. Results are presentative of at least 3 independent experiments (A–E). (F) Bar graphs were obtained by quantification of ATF6 nuclear staining (*P < 0.05 versus control). Results are presentative of 3 independent experiments (A–E) or the mean ± S.E.M. of an experimental n = 3 (F). Image collected & cropped by CiteAb from the following publication (https://www.oncotarget.com/lookup/doi/10.18632/oncotarget.15469), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

Immunocytochemistry/ Immunofluorescence: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] -

Immunocytochemistry/ Immunofluorescence: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - UPR status in STC-1 & GluTag cell lines & effect of UPR inducers on markers of the UPR pathwaysSTC-1 & GluTag cells were incubated in medium (Ctrl) or ER stress-inducing agents thapsigargin (Tg, 300 nM), tunicamycin (Tn, 0.05 μg/mL) & brefeldin A (Bref A, 3 μM) for 4 h, 16 h & 8 h respectively. (A) Protein expression level of phosphorylated or total forms of PERK, eIF2 alpha & CHOP, BiP & cleaved-caspase 3 (C-Caspase 3) protein expression was examined using Western Blot analysis. alpha -tubulin was used as internal control (B) Densitometric quantification of P-PERK/PERK, P-eIF2 alpha /eIF2 alpha & BiP/ alpha -tubulin ratios analysis in STC-1 or GluTag cell lines (*P < 0.05 versus control). (C–D) The effect of Tg, Tn & Bref A on ATF6 nuclear localization was assessed by immunofluorescence in STC-1 cells (C) & GluTag cells (D) using anti-ATF6 antibody & Hoechst dye. Magnification ×1000. (E) Bar graphs obtained by quantification of ATF6 nuclear staining (*P < 0.05 versus control). (F) XBP1 mRNA splicing was analyzed by RT-PCR after Pst1 digestion: XBP1-h, hybrid, XBP1-u, unspliced; XBP1-s, spliced variant of XBP1; *, XBP1-u mRNA fragments after Pst1 digestion. Results are presentative of 3 independent experiments (A, C, D, F) or the mean ± S.E.M. of an experimental n = 3 (B, E). Image collected & cropped by CiteAb from the following publication (https://www.oncotarget.com/lookup/doi/10.18632/oncotarget.15469), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

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

Western Blot: ATF6 Antibody (70B1413.1) - BSA Free [NBP1-40256] - Tm‐induced UPR increases expression of RANKL in cultured osteoblastic & osteocytic cells. (A) Western blotting of cell lysates obtained from neonatal calvaria‐derived osteoblastic cells (Calvaria Ob) treated with 2.2 µg/mL Tm for the indicated times. (B‐D) Gene expression as determined by qRT‐PCR in (B) calvaria‐derived osteoblastic cells (n = 3/group) (C) Osteoblastic UAMS‐32 cells (UAMS‐32 Ob) (n = 4/group) or (D) osteocytic MLO‐Y4 cells (MLO‐Y4 Ot) (n = 4/group),maintained in presence of vehicle (, 0.1% DMSO) or 2.2 µg/ml Tm () for 4 hours. (E) Western blot of RANKL protein in cell lysates obtained from calvaria‐derived osteoblastic cells as described in A in a separate study. Data shown are the mean & SD with individual data points. *P < .05 vs vehicle by Student's t‐test Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32259048), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
ATF6 Antibody (70B1413.1) - BSA Free

Immunocytochemistry/Immunofluorescence: Mouse Monoclonal ATF6 Antibody (70B1413.1) [IMGENEX: IMG-273] [NBP1-40256]

Immunocytochemistry/Immunofluorescence: Mouse Monoclonal ATF6 Antibody (70B1413.1) [IMGENEX: IMG-273] [NBP1-40256] - Mice hepatocytes were stained for ATF6. Image from a verified customer review.
ATF6 Antibody (70B1413.1) - BSA Free

Western Blot: Mouse Monoclonal ATF6 Antibody (70B1413.1) [IMGENEX: IMG-273] [NBP1-40256]

Western Blot: Mouse Monoclonal ATF6 Antibody (70B1413.1) [IMGENEX: IMG-273] [NBP1-40256] - Western blot of the mouse liver homogenates from control (1) and alcohol-fed mice (2). Image from a verified customer review.

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 7 reviews rated 4.1 using NBP1-40256 in the following applications:

Flow Cytometry Panel Builder

Bio-Techne Knows Flow Cytometry

Save time and reduce costly mistakes by quickly finding compatible reagents using the Panel Builder Tool.

Advanced Features

  • Spectra Viewer - Custom analysis of spectra from multiple fluorochromes
  • Spillover Popups - Visualize the spectra of individual fluorochromes
  • Antigen Density Selector - Match fluorochrome brightness with antigen density
Build Your Panel Now
Flow Cytometry

Formulation, Preparation, and Storage

Purification

Protein G purified

Formulation

PBS

Format

BSA Free

Preservative

0.02% 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

P18850

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 or batch number in the search box below.

Download SDS

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.

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

Customer Reviews for ATF6 Antibody (70B1413.1) - BSA Free (7)

4.1 out of 5
7 Customer Ratings
5 Stars
57%
4 Stars
29%
3 Stars
0%
2 Stars
0%
1 Stars
14%

Have you used ATF6 Antibody (70B1413.1) - BSA Free?

Submit a review and receive an Amazon gift card!

$25/€18/£15/$25CAN/¥2500 Yen for a review with an image

$10/€7/£6/$10CAN/¥1110 Yen for a review without an image

Submit a review
Amazon Gift Card

Customer Images

Showing  1 - 5 of 7 reviews Showing All
Filter By:
Clear Filters X
  • ATF6 Antibody (70B1413.1) - BSA Free
    Name: Armen Petrosyan
    Application: Western Blot
    Sample Tested: Liver homogenates sample
    Species: Mouse
    Verified Customer | Posted 11/19/2024
    W-B of the mouse liver homogenates from control (1) and alcohol-fed mice (2).
    ATF6 Antibody (70B1413.1) - BSA Free NBP1-40256
  • ATF6 Antibody (70B1413.1) - BSA Free
    Name: Armen Petrosyan
    Application: Immunofluorescence
    Sample Tested: Primary mouse hepatocytes
    Species: Mouse
    Verified Customer | Posted 11/19/2024
    Mice hepatocytes were stained for NBP1-40256
    ATF6 Antibody (70B1413.1) - BSA Free NBP1-40256
  • ATF6 Antibody (70B1413.1) - BSA Free
    Name: Armen Petrosyan
    Application: Western Blot
    Sample Tested: PC-3 human prostate cancer cell line
    Species: Human
    Verified Customer | Posted 06/27/2023
    Lysate of PC-3 cells
    ATF6 Antibody (70B1413.1) - BSA Free NBP1-40256
  • ATF6 Antibody (70B1413.1) - BSA Free
    Name: Anonymous
    Application: Western Blot
    Sample Tested: DU145 human prostate carcinoma cell line
    Species: Human
    Verified Customer | Posted 06/27/2023
    Lysate of DU145 cells
    ATF6 Antibody (70B1413.1) - BSA Free NBP1-40256
  • ATF6 Antibody (70B1413.1)
    Name: Anonymous
    Application: Immunohistochemistry-Paraffin
    Sample Tested: Placental tissue
    Species: Human
    Verified Customer | Posted 01/24/2022
    Placenta staining
    Fixed paraffin-embedded sections. Antibody dilution 1:50. Incubated overnight in 4C.
    ATF6 Antibody (70B1413.1) - BSA Free NBP1-40256
  • ATF6 Antibody (70B1413.1)
    Name: yann Jaudouin
    Application: WES protein simple
    Sample Tested: cultured embryonic cortical neuron lysate 0
    Species: Rat
    Verified Customer | Posted 09/14/2018
    ATF6 1/25e
    Lysate protein cortical neuron 0,2 mg/ml. test antibody: 1/25e, 1/50e, 1/100e
    ATF6 Antibody (70B1413.1) - BSA Free NBP1-40256
  • Name: Shinobu Miyazaki
    Application: Western Blot
    Sample Tested: Whole cell lysate
    Species: Mouse
    Verified Customer | Posted 03/18/2016
    ATF6 antibody review
    ATF6 Antibody (70B1413.1) - BSA Free NBP1-40256

There are no reviews that match your criteria.

Showing  1 - 5 of 7 reviews Showing All

Protocols

View specific protocols for ATF6 Antibody (70B1413.1) - BSA Free (NBP1-40256):

Protocol for Flow Cytometry Intracellular Staining
Sample Preparation.
1. Grow cells to 60-85% confluency. Flow cytometry requires between 2 x 105 and 1 x 106 cells for optimal performance.
2. If cells are adherent, harvest gently by washing once with staining buffer and then scraping. Avoid using trypsin as this can disrupt certain epitopes of interest. If enzymatic harvest is required, use Accutase, Collagenase, or TrypLE Express for a less damaging option.
3. Reserve 100 uL for counting, then transfer cell volume into a 50 mL conical tube and centrifuge for 8 minutes at 400 RCF.
a. Count cells using a hemocytometer and a 1:1 trypan blue exclusion stain to determine cell viability before starting the flow protocol. If cells appear blue, do not proceed.
4. Re-suspend cells to a concentration of 1 x 106 cells/mL in staining buffer (NBP2-26247).
5. Aliquot out 1 mL samples in accordance with your experimental samples.

Tip: When cell surface and intracellular staining are required in the same sample, it is advisable that the cell surface staining be performed first since the fixation and permeablization steps might reduce the availability of surface antigens.

Intracellular Staining.
Tip: When performing intracellular staining, it is important to use appropriate fixation and permeabilization reagents based upon the target and its subcellular location. Generally, our Intracellular Flow Assay Kit (NBP2-29450) is a good place to start as it contains an optimized combination of reagents for intracellular staining as well as an inhibitor of intracellular protein transport (necessary if staining secreted proteins). Certain targets may require more gentle or transient permeabilization protocols such as the commonly employed methanol or saponin-based methods.
Protocol for Cytoplasmic Targets:
Optional: Perform cell surface staining as described in the previous section.
1. Fix the cells by adding 100 uL fixation solution (such as 4% PFA) to each sample for 10-15 minutes.
2. Permeabilize cells by adding 100 uL of a permeabization buffer to every 1 x 106 cells present in the sample. Mix well and incubate at room temperature for 15 minutes.
a. For cytoplasmic targets, use a gentle permeabilization solution such as 1X PBS + 0.5% Saponin or 1X PBS + 0.5% Tween-20.
b. To maintain the permeabilized state throughout your experiment, use staining buffer + 0.1% of the permeabilization reagent (i.e. 0.1% Tween-20 or 0.1% Saponin).
3. Following the 15 minute incubation, add 2 mL of the staining buffer + 0.1% permeabilizer to each sample.
4. Centrifuge for 5 minutes at 400 RCF.
5. Discard supernatant and re-suspend in 1 mL of staining buffer + 0.1% permeabilizer.
6. Stain each sample at 1 uL/ 1 x 106 cells of primary antibody or 1-3 uL/ 1 x 106 cells for directly conjugated antibodies. Mix well and incubate at room temperature for 30 minutes- 1 hour. Gently mix samples every 10-15 minutes.
7. Following the primary/conjugate incubation, add 2 mL/sample of staining buffer +0.1% permeabilizer and centrifuge for 5 minutes at 400 RCF.
8. Remove supernatant and re-suspend each sample in 2 mL staining buffer + 0.1% permeabilizer, repeat wash for 5 minutes at 400 RCF.
9. If using a directly conjugated antibody, after the second wash, re-suspend cell pellet to a final volume of 500 uL per sample and proceed with flow analysis.

Immunocytochemistry Protocol

Culture cells to appropriate density in 35 mm culture dishes or 6-well plates.

1. Remove culture medium and wash the cells briefly in PBS. Add 10% formalin to the dish and fix at room temperature for 10 minutes.
2. Remove the formalin and wash the cells in PBS.
3. Permeablize the cells with 0.1% Triton X100 or other suitable detergent for 10 min.
4. Remove the permeablization buffer and wash three times for 10 minutes each in PBS. Be sure to not let the specimen dry out.
5. To block nonspecific antibody binding, incubate in 10% normal goat serum from 1 hour to overnight at room temperature.
6. Add primary antibody at appropriate dilution and incubate overnight at 4C.
7. Remove primary antibody and replace with PBS. Wash three times for 10 minutes each.
8. Add secondary antibody at appropriate dilution. Incubate for 1 hour at room temperature.
9. Remove secondary antibody and replace with PBS. Wash three times for 10 minutes each.
10. Counter stain DNA with DAPi if required.

Immunohistochemistry-Paraffin Embedded Sections

Antigen Unmasking:
Bring slides to a boil in 10 mM sodium citrate buffer (pH 6.0) then maintain at a sub-boiling temperature for 10 minutes. Cool slides on bench-top for 30 minutes (keep slides in the sodium citrate buffer at all times).

Staining:
1. Wash sections in deionized water three times for 5 minutes each.
2. Wash sections in PBS for 5 minutes.
3. Block each section with 100-400 ul blocking solution (1% BSA in PBS) for 1 hour at room temperature.
4. Remove blocking solution and add 100-400 ul diluted primary antibody. Incubate overnight at 4 C.
5. Remove antibody solution and wash sections in wash buffer three times for 5 minutes each.
6. Add 100-400 ul HRP polymer conjugated secondary antibody. Incubate 30 minutes at room temperature.
7. Wash sections three times in wash buffer for 5 minutes each.
8. Add 100-400 ul DAB substrate to each section and monitor staining closely.
9. As soon as the sections develop, immerse slides in deionized water.
10. Counterstain sections in hematoxylin.
11. Wash sections in deionized water two times for 5 minutes each.
12. Dehydrate sections.
13. Mount coverslips.


Western Blot Protocol

1. Perform SDS-PAGE on samples to be analyzed, loading 10-25 ug of total protein per lane.
2. Transfer proteins to PVDF membrane according to the instructions provided by the manufacturer of the membrane and transfer apparatus.
3. Stain the membrane with Ponceau S (or similar product) to assess transfer success, and mark molecular weight standards where appropriate.
4. Rinse the blot TBS -0.05% Tween 20 (TBST).
5. Block the membrane in 5% Non-fat milk in TBST (blocking buffer) for at least 1 hour.
6. Wash the membrane in TBST three times for 10 minutes each.
7. Dilute primary antibody in blocking buffer and incubate overnight at 4C with gentle rocking.
8. Wash the membrane in TBST three times for 10 minutes each.
9. Incubate the membrane in diluted HRP conjugated secondary antibody in blocking buffer (as per manufacturer's instructions) for 1 hour at room temperature.
10. Wash the blot in TBST three times for 10 minutes each (this step can be repeated as required to reduce background).
11. Apply the detection reagent of choice in accordance with the manufacturer's instructions.

Find general support by application which include: protocols, troubleshooting, illustrated assays, videos and webinars.

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

Showing  1 - 5 of 5 FAQs Showing All

    • Q: I learn from your company web that you are providing atf6, xbp1, cbx4 antibodies against human for immunofluorescence. And from the provided pictures, it seems very good. But I am wondering if anyone use them for immunofluorescence in human cells in your memory or in published papers? And how about them? By the way, you are providing free sample size for the atf6, xbp1 antibodies, aren't you? Could you send me some for tests? One more question, do they work on endogenous proteins but not only on overexpressed proteins?
      A: These products have been validated for use in IF on human: catalog numbers NBP1-40256, NB100-80861, and NBP1-83225. Yes these antibodies should work on the native protein/endogenous protein.

    • A: This antibody was made against a partial recombinant protein (not a short peptide). As such we do not have a blocking peptide for this antibody nor do we have a blocking protein. We do however have a positive control, ready to use ATF6 lysate NBP1-70778.

    • A: NBP1-40256 (ATF6 Antibody/70B1413.1) can be used in WB with mouse tissue/cell line samples. We have many secondary antibodies that can potentially be used with NBP1-40256. However, since the primary antibody has an isotype of IgG1, it is better to use the WB-validated anti-mouse IgG1 secondary antibodies.

    • Q: We are having trouble with the following antibody, we did not observe any signal for ATF6 (full length or cleaved), only a non-specific band at around 37 kDa. We've tested on human MRC-5 fibrablasts. Do you have any suggestions that may be helpful to us? 
      A: The nature of the protein target ATF6 is differently expressed as the unglycosylated and/or glycosylated forms, with the different levels of glycosylation (http://www.uniprot.org/uniprot/P18850#ptm_processing), in the different organs/tissues/cells under the different physiological/pathological conditions. This ATF6 Antibody/70B1413.1 is a mouse monoclonal antibody with a very narrow epitope (even though we don't map any epitopes for our antibodies, it is known that almost all monoclonal epitopes have the sizes less than 15 a.a.). In other words, NBP1-40256 can bind to the full-length ATF6 and the long fragment of the cleaved protein target (not both cleaved protein species, and see #3 below). In order to detect the full-length and both two cleaved fragments of ATF6, the immunogen has to extend before and after the cleavage site at a.a. 419/420, in both directions, and therefore the epitopes can present in both sides of the cleavage site.  ATF6 has 1 transmembrane domain, 1 leucine-zipper, and three glycosylation sites that could affect the mobility in electrophoresis. As the rule of thumb, we use 110 Da as an average of one amino acid. Therefore the full-length ATF6 gives 73,700 Da (vs. 74,585 Da calculated by the exact sequence). Since it is cleaved at a.a. 419 of ATF6, two fragments will be 419 a.a. and 251 a.a. (670 - 419). Take each of them and times 110 Da individually, and these are the results: 46,090 Da and 27,610 Da (the estimated values), respectively.  Thefore your ~37 kDa species might be the short protein species with glycosylations (please double check with #1 above). Finally, we would like to suggest two things: please include other human cell lines as the controls: http://www.proteinatlas.org/ENSG00000118217-ATF6/antibody shows the WB positive results derived from using two cell lines and one organ. These could be used as the positive control. Also, the glycosylation levels are likely to be different in the different cells/tissues/organs, and therefor the different sizes of ATF can be viewed.  The second thing is to use a real positive control lysate, such as ATF6 cell lysate or ATF6 overexpression lysate for the specificity of NBP1-40256.

    • Q: We are looking for an antibody specific to ATF6 alpha that will not cross-react with ATF6 beta. Would you please help confirm if any of the following are ATF6 alpha specific antibodies: NBP1-40256, NBP1-77251, NBP1-76675 & NBP1-41439?
      A: The antibodies NBP1-77251 and NBP1-76675 are specific to ATF6 alpha and should not cross-react with ATF6 beta.
Showing  1 - 5 of 5 FAQs Showing All
View all FAQs for Antibodies