Key Product Details

Validated by

Biological Validation

Species Reactivity

Validated:

Human, Mouse, Rat

Cited:

Human, Mouse, Rat, Hamster

Applications

Validated:

Western Blot, Immunocytochemistry, Simple Western

Cited:

Immunohistochemistry, Immunohistochemistry-Frozen, Western Blot, Immunocytochemistry, Co-Immunoprecipitation, ELISA Capture

Label

Unconjugated

Antibody Source

Polyclonal Rabbit IgG
View all HTRA2/Omi Primary Antibodies »

Product Specifications

Immunogen

E. coli-derived recombinant human HTRA2/Omi
Ala134-Glu458
Accession # O43464

Specificity

Detects human, mouse, and rat full length and mitochondria-processed HTRA2/Omi.

Clonality

Polyclonal

Host

Rabbit

Isotype

IgG

Scientific Data Images for Human/Mouse/Rat HTRA2/Omi Antibody

Detection of Human/Mouse/Rat HTRA2/Omi antibody by Western Blot.

Detection of Human/Mouse/Rat HTRA2/Omi by Western Blot.

Western blot shows lysates of PC-12 rat adrenal pheochromocytoma cell line, Jurkat human acute T cell leukemia cell line, HeLa human cervical epithelial carcinoma cell line, L-929 mouse fibroblast cell line, and C2C12 mouse myoblast cell line. PVDF membrane was probed with 0.25 µg/mL of Rabbit Anti-Human/Mouse/Rat HTRA2/Omi Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1458) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). Specific bands were detected for HTRA2/Omi at approximately 36 and 49 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 2.
HTRA2/Omi antibody in Jurkat Human Cell Line by Immunocytochemistry (ICC).

HTRA2/Omi in Jurkat Human Cell Line.

HTRA2/Omi was detected in immersion fixed Jurkat human acute T cell leukemia cell line stimulated with staurosporin using Rabbit Anti-Human/Mouse/Rat HTRA2/Omi Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1458) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Rabbit IgG Secondary Antibody (yellow; Catalog # NL004) and counterstained with DAPI (blue). View our protocol for Fluorescent ICC Staining of Cells on Coverslips.
Detection of Human and Mouse HTRA2/Omi antibody by Simple WesternTM.

Detection of Human and Mouse HTRA2/Omi by Simple WesternTM.

Simple Western lane view shows lysates of C2C12 mouse myoblast cell line and HeLa human cervical epithelial carcinoma cell line, loaded at 0.2 mg/mL. Specific bands were detected for HTRA2/Omi at approximately 51 kDa (precursor) and 41 kDa (processed) (as indicated) using 2.5 µg/mL of Rabbit Anti-Human/Mouse/Rat HTRA2/Omi Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1458). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.
Detection of Human HTRA2/Omi by Western Blot

Detection of Human HTRA2/Omi by Western Blot

Caspases are not activated by FGFR inhibition in FGFR2‐mutant EC cells. (A) Western blots showing total caspase‐3 and caspase‐7 in response to treatment with 300 nm BGJ398 for up to 72 h. Tubulin serves as a loading control. *Denotes nonspecific band. (B) AN3CA and JHUEM2 cells were pretreated with 100 μm Z‐VAD‐FMK for 1 h prior to the addition of DMSO, 1 μm PD173074 (PD), 300 nm BGJ398 (BGJ) or 300 nm AZD4547 (AZD) for 72 h. Cell death was detected by staining cells with Annexin V. The mean percentage of Annexin V‐positive cells from three independent experiments (each performed in triplicate) is shown along with SD. (C) Western blot showing cleavage of caspase‐3 in AN3CA and JHUEM2 cells treated with 1 μm actinomycin D (Act D) or staurosporine (STS), respectively, for 24 h. (D) Mean percentage of AN3CA and JHUEM2 cells showing Annexin V‐positive staining following treatment with 100 μm Z‐VAD‐FMK alone or 1 h prior to treatment with 1 μm Act D and STS for 24 h. The mean from three independent experiments (each performed in triplicate) is shown along with SD. (E) Western blot showing staining of Bim, Bid, HTRA2/OMI and Smac/diablo in cytosolic and mitochondrial fractions of JHUEM2 cells treated with 300 nm BGJ398 for 24 and 48 h. Tom20 serves as a marker of the mitochondrial fraction, GRP78 as a marker of the ER, Lamin B1 as a marker of the nuclear fraction and GAPDH as a marker of the cytosolic fraction. (F) Western blot showing staining of AIF in mitochondrial and nuclear fractions of AN3CA cells treated with 1 μm PD173074 for 48 h. Cox IV and PARP serve as mitochondrial and nuclear markers, respectively. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/30537101), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Mouse HTRA2/Omi by Western Blot

Detection of Mouse HTRA2/Omi by Western Blot

Neural deletion of Htra2 is sufficient to generate neurological phenotypes.(A) Exons 2 to 4 of Htra2 were flanked with loxP sites, with a FRT flanked neo cassette 3′ to exon 4. Expression of FlpE causes deletion of the selection cassette. Cre-mediated deletion causes excision of exons 2 to 4. Small arrows beneath the allele constructs denote the position of genotyping primers. (B) PCR from genomic DNA can distinguish WT (+, arrow, 279 bp), KO (–, filled arrowhead, 358 bp) and floxed (f, empty arrowhead, 313 bp) alleles of Htra2. (C) Western blot analysis confirmed loss of HTRA2 protein (arrow) in all tissues of HTRA2 KO mice and reduction in brain of NesKO mice (arrowheads denote non-specific bands). The levels of HTRA2 protein in NesKO spleen and thymus were comparable with NesWT. Cx: cortex, Mb: midbrain, Hb: hindbrain. PHB2 was used as a loading control. (D) HTRA2 KO mice and NesKO mice were smaller than WT littermates by comparison. The size of the thymus and spleen was reduced although brain was relatively normal in size (representative animals shown at P30, scale bar: 1 cm.). (E) Body weight of HTRA2 KO and NesKO mice did not increase beyond P18 (n = 56 (HTRA2 WT), 62 (HTRA2 KO), 35 (NesWT), 25 (NesKO), error bars indicate SEM). Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/25531304), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Mouse HTRA2/Omi by Western Blot

Detection of Mouse HTRA2/Omi by Western Blot

Neural deletion of Htra2 is sufficient to generate neurological phenotypes.(A) Exons 2 to 4 of Htra2 were flanked with loxP sites, with a FRT flanked neo cassette 3′ to exon 4. Expression of FlpE causes deletion of the selection cassette. Cre-mediated deletion causes excision of exons 2 to 4. Small arrows beneath the allele constructs denote the position of genotyping primers. (B) PCR from genomic DNA can distinguish WT (+, arrow, 279 bp), KO (–, filled arrowhead, 358 bp) and floxed (f, empty arrowhead, 313 bp) alleles of Htra2. (C) Western blot analysis confirmed loss of HTRA2 protein (arrow) in all tissues of HTRA2 KO mice and reduction in brain of NesKO mice (arrowheads denote non-specific bands). The levels of HTRA2 protein in NesKO spleen and thymus were comparable with NesWT. Cx: cortex, Mb: midbrain, Hb: hindbrain. PHB2 was used as a loading control. (D) HTRA2 KO mice and NesKO mice were smaller than WT littermates by comparison. The size of the thymus and spleen was reduced although brain was relatively normal in size (representative animals shown at P30, scale bar: 1 cm.). (E) Body weight of HTRA2 KO and NesKO mice did not increase beyond P18 (n = 56 (HTRA2 WT), 62 (HTRA2 KO), 35 (NesWT), 25 (NesKO), error bars indicate SEM). Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/25531304), licensed under a CC-BY license. Not internally tested by R&D Systems.

Applications for Human/Mouse/Rat HTRA2/Omi Antibody

Application
Recommended Usage

Immunocytochemistry

5-15 µg/mL
Sample: Immersion fixed non-apoptotic Jurkat human acute T cell leukemia cell line and Staurosporine treated apoptotic Jurkat human acute T cell leukemia cell line

Simple Western

2.5 µg/mL
Sample: C2C12 mouse myoblast cell line and HeLa human cervical epithelial carcinoma cell line

Western Blot

0.25 µg/mL
Sample: PC-12 rat adrenal pheochromocytoma cell line, Jurkat human acute T cell leukemia cell line, HeLa human cervical epithelial carcinoma cell line, L-929 mouse fibroblast cell line, and C2C12 mouse myoblast cell line

Reviewed Applications

Read 3 reviews rated 4.3 using AF1458 in the following applications:

Formulation, Preparation, and Storage

Purification

Antigen Affinity-purified

Reconstitution

Reconstitute at 0.2 mg/mL in sterile PBS. For liquid material, refer to CoA for concentration.

Formulation

Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.

Shipping

Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.

Stability & Storage

Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.

Calculators

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: HTRA2/Omi

HtrA2/Omi is the mammalian homologue of bacterial high temperature requirement protein (HtrA). HtrA2/Omi localizes to the mitochondria and is processed to expose an amino-terminal Reaper-like motif similar to SMAC/Diablo. HtrA2/Omi is released from the mitochondria in response to apoptotic insult and can interact with the BIR2 or BIR3 domains of XIAP to relieve caspase-IAP inhibition. This effect can be measured by reversing XIAP-BIR2 (R&D Systems, Catalog # 786-XB) inhibition of Caspase-7 (R&D Systems, Catalog # 823-C7) cleavage of a fluorogenic peptide (DEVD-AFC, MP Bio, Catalog # AFC-138). IC50 values for this effect are typically between 0.2 and 1.5 μM. HtrA2/Omi is trimeric and functions as a serine protease. The serine protease activity may play a more central role in apoptosis than its IAP antagonizing function. A PDZ domain regulates the serine protease activity by blocking access to the active site. The specificity of the protease is yet to be defined and no endogenous substrates are known to date.

References

  1. Suzuki, Y. et al. (2001) Mol. Cell. 8:613.
  2. van Loo, G. et al. (2002) Cell Death & Diff. 9:20.
  3. Hedge, R. et al. (2001) J. Biol. Chem. 277:432.
  4. Verhagen, A. et al. (2001) J. Biol. Chem. 277:445.
  5. Martins, L. et al. (2002) J. Biol. Chem. 277:439.
  6. Silke, J., and A. Verhagen (2002) Cell Death & Diff. 9:362.
  7. Savopoulos, J. et al. (2000) Protein Expression & Purification 19:227.

Long Name

High Temperature Requirement Protein-2

Alternate Names

Omi, PRSS25

Entrez Gene IDs

27429 (Human); 64704 (Mouse)

Gene Symbol

HTRA2

UniProt

O43464

Additional HTRA2/Omi Products

Product Documents for Human/Mouse/Rat HTRA2/Omi Antibody

Certificate of Analysis

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Product Specific Notices for Human/Mouse/Rat HTRA2/Omi Antibody

For research use only

Citations for Human/Mouse/Rat HTRA2/Omi Antibody

Customer Reviews for Human/Mouse/Rat HTRA2/Omi Antibody (3)

4.3 out of 5
3 Customer Ratings
5 Stars
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Showing  1 - 3 of 3 reviews Showing All
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  • Human/Mouse/Rat HTRA2/Omi Antibody
    Name: Anonymous
    Application: Immunocytochemistry
    Sample Tested: pig SC cells
    Species: Pig
    Verified Customer | Posted 06/07/2017
    AF1458 HTRA2 ab labeled pig SC cells with Alexa594
    Human/Mouse/Rat HTRA2/Omi Antibody AF1458
  • Human/Mouse/Rat HTRA2/Omi Antibody
    Name: Anonymous
    Application: Immunocytochemistry
    Sample Tested: Hep2G cells
    Species: Pig
    Verified Customer | Posted 06/07/2017
    AF1458, HTRA2 antibody labeling Hep2G cells with Alexa594
    Human/Mouse/Rat HTRA2/Omi Antibody AF1458
  • Name: Anonymous
    Application: Immunofluorescence
    Sample Tested: See PMID 23183826
    Species: Human
    Verified Customer | Posted 01/05/2015

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Associated Pathways

Apoptosis Signaling Pathway Apoptosis Signaling Pathway Thumbnail