HTRA2/Omi Antibody (229926) [Unconjugated]

Western blot shows lysates of SK-Mel-28 human malignant melanoma cell line, MCF-7 human breast cancer cell line, HEK293 human embryonic kidney cell line, and HeLa human cervical epithelial carcinoma cell line. PVDF membrane was probed with 1 µg/mL of Mouse Anti-Human HTRA2/Omi Monoclonal Antibody (Catalog # MAB1458) followed by HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # HAF007). A specific band was detected for HTRA2/Omi at approximately 35 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 4.

HTRA2/Omi was detected in immersion fixed K562 human chronic myelogenous leukemia cell line using Mouse Anti-Human HTRA2/Omi Monoclonal Antibody (Catalog # MAB1458) at 25 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Mouse IgG Secondary Antibody (red; NL007) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm. Staining was performed using our protocol for Fluorescent ICC Staining of Non-adherent Cells.

HtrA2/Omi expression increases following CMV infection.(A–B) Immunoblot analyses of HtrA2/Omi in lysates of mock-infected HF (M) or HF infected (MOI of 3) with Towne-BAC or delta UL37x1 for 24, 48, 72, 96, or 120 h and control cell lysates from HeLa cells transfected with HtrA2/Omi expression plasmid (+) or control plasmid (−). The expected migration of the 49 kDa immature, pro-HtrA2/Omi and 36 kDA mature, HtrA2/Omi was calculated based on migration of molecular weight markers (not shown). Immunoblot detection of HtrA2, beta -actin and IE1 are shown. (C–N) Immunofluorescent images of HtrA2/Omi (C, F) and cytochrome c (I, L) (red), and GFP fluorescence (green) (D, G, J, M) and merged images (E, H, K, N) at 72 h (C–H) or 96 h (I–N) postinfection (MOI of 0.001) with delta UL37x1 (C–E, I–K) or Towne-BAC (F–H, L–N). Original magnification ×1000. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/18769594), licensed under a CC-BY license. Not internally tested by R&D Systems.

HtrA2/Omi expression increases following CMV infection.(A–B) Immunoblot analyses of HtrA2/Omi in lysates of mock-infected HF (M) or HF infected (MOI of 3) with Towne-BAC or delta UL37x1 for 24, 48, 72, 96, or 120 h and control cell lysates from HeLa cells transfected with HtrA2/Omi expression plasmid (+) or control plasmid (−). The expected migration of the 49 kDa immature, pro-HtrA2/Omi and 36 kDA mature, HtrA2/Omi was calculated based on migration of molecular weight markers (not shown). Immunoblot detection of HtrA2, beta -actin and IE1 are shown. (C–N) Immunofluorescent images of HtrA2/Omi (C, F) and cytochrome c (I, L) (red), and GFP fluorescence (green) (D, G, J, M) and merged images (E, H, K, N) at 72 h (C–H) or 96 h (I–N) postinfection (MOI of 0.001) with delta UL37x1 (C–E, I–K) or Towne-BAC (F–H, L–N). Original magnification ×1000. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/18769594), licensed under a CC-BY license. Not internally tested by R&D Systems.

Transient overexpression of HtrA2/Omi induces early death in CMV-infected but not uninfected cells.(A–F) Immunofluorescent images of HtrA2/Omi (A–C) and GFP fluorescence (D–F) following cotransfection of HF with GFP expression plasmid together with empty vector, HtrA2/Omi, or HtrA2S306A expression plasmids. Original magnification ×400. (G) Immunoblot depicting 49 kDa, immature and 36 kDA, mature forms of HtrA2/Omi and HtrA2S306A as well as beta -actin control in transfected HeLa cell lysates. (H) Percentage of live HFs at 120 h post cotransfection of GFP expression plasmid with vector, HtrA2/Omi, or HtrA2S306A expression plasmids. (I) Number of viral plaques 10 days following cotransfection of delta UL37x1 or Towne-BAC DNA (500 ng) with 800 ng vector, HtrA2/Omi, or HtrA2S306A expression plasmids. (J) Number of live (intact) cells in cultures at 48 or 72 h following transfection with Towne-BAC DNA (300 ng) alone or with 333 ng of HtrA2 or HtrA2S306A expression plasmids. (K) Number of live (intact) cells (±range) at 48 or 72 h post transfection with Towne-BAC DNA (500 ng) and 300 ng of vector or HtrA2/Omi expression plasmid with or without addition of 10 µM UCF-101 from 6 h. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/18769594), licensed under a CC-BY license. Not internally tested by R&D Systems.

HtrA2/Omi expression increases following CMV infection.(A–B) Immunoblot analyses of HtrA2/Omi in lysates of mock-infected HF (M) or HF infected (MOI of 3) with Towne-BAC or delta UL37x1 for 24, 48, 72, 96, or 120 h and control cell lysates from HeLa cells transfected with HtrA2/Omi expression plasmid (+) or control plasmid (−). The expected migration of the 49 kDa immature, pro-HtrA2/Omi and 36 kDA mature, HtrA2/Omi was calculated based on migration of molecular weight markers (not shown). Immunoblot detection of HtrA2, beta -actin and IE1 are shown. (C–N) Immunofluorescent images of HtrA2/Omi (C, F) and cytochrome c (I, L) (red), and GFP fluorescence (green) (D, G, J, M) and merged images (E, H, K, N) at 72 h (C–H) or 96 h (I–N) postinfection (MOI of 0.001) with delta UL37x1 (C–E, I–K) or Towne-BAC (F–H, L–N). Original magnification ×1000. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/18769594), licensed under a CC-BY license. Not internally tested by R&D Systems.

Transient overexpression of HtrA2/Omi induces early death in CMV-infected but not uninfected cells.(A–F) Immunofluorescent images of HtrA2/Omi (A–C) and GFP fluorescence (D–F) following cotransfection of HF with GFP expression plasmid together with empty vector, HtrA2/Omi, or HtrA2S306A expression plasmids. Original magnification ×400. (G) Immunoblot depicting 49 kDa, immature and 36 kDA, mature forms of HtrA2/Omi and HtrA2S306A as well as beta -actin control in transfected HeLa cell lysates. (H) Percentage of live HFs at 120 h post cotransfection of GFP expression plasmid with vector, HtrA2/Omi, or HtrA2S306A expression plasmids. (I) Number of viral plaques 10 days following cotransfection of delta UL37x1 or Towne-BAC DNA (500 ng) with 800 ng vector, HtrA2/Omi, or HtrA2S306A expression plasmids. (J) Number of live (intact) cells in cultures at 48 or 72 h following transfection with Towne-BAC DNA (300 ng) alone or with 333 ng of HtrA2 or HtrA2S306A expression plasmids. (K) Number of live (intact) cells (±range) at 48 or 72 h post transfection with Towne-BAC DNA (500 ng) and 300 ng of vector or HtrA2/Omi expression plasmid with or without addition of 10 µM UCF-101 from 6 h. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/18769594), licensed under a CC-BY license. Not internally tested by R&D Systems.

Transient overexpression of HtrA2/Omi induces early death in CMV-infected but not uninfected cells.(A–F) Immunofluorescent images of HtrA2/Omi (A–C) and GFP fluorescence (D–F) following cotransfection of HF with GFP expression plasmid together with empty vector, HtrA2/Omi, or HtrA2S306A expression plasmids. Original magnification ×400. (G) Immunoblot depicting 49 kDa, immature and 36 kDA, mature forms of HtrA2/Omi and HtrA2S306A as well as beta -actin control in transfected HeLa cell lysates. (H) Percentage of live HFs at 120 h post cotransfection of GFP expression plasmid with vector, HtrA2/Omi, or HtrA2S306A expression plasmids. (I) Number of viral plaques 10 days following cotransfection of delta UL37x1 or Towne-BAC DNA (500 ng) with 800 ng vector, HtrA2/Omi, or HtrA2S306A expression plasmids. (J) Number of live (intact) cells in cultures at 48 or 72 h following transfection with Towne-BAC DNA (300 ng) alone or with 333 ng of HtrA2 or HtrA2S306A expression plasmids. (K) Number of live (intact) cells (±range) at 48 or 72 h post transfection with Towne-BAC DNA (500 ng) and 300 ng of vector or HtrA2/Omi expression plasmid with or without addition of 10 µM UCF-101 from 6 h. Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/18769594), licensed under a CC-BY license. Not internally tested by R&D Systems.

• Reactivity: Hu
• Applications: WB, ICC/IF
• Clone: 229926
• Clonality: Monoclonal
• Host: Mouse
• Conjugate: Unconjugated
• Concentration: LYOPH

HTRA2/Omi Antibody (229926) [Unconjugated] Summary

• Immunogen: E. coli-derived recombinant human HTRA2/Omi
  Ala134-Glu458
  Accession # O43464
• Specificity: Detects the mitochondria-processed form of human HTRA2/Omi.
• Source: N/A
• Isotype: IgG2b
• Clonality: Monoclonal
• Host: Mouse
• Gene: HTRA2
• Purity Statement: Protein A or G purified from hybridoma culture supernatant

Applications/Dilutions

• Dilutions:
  • Immunocytochemistry 8-25 ug/mL
  • Western Blot 1 ug/mL

Packaging, Storage & Formulations

• 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.
• Buffer: 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.
• Preservative: No Preservative
• Concentration: LYOPH
• Reconstitution Instructions: Reconstitute at 0.5 mg/mL in sterile PBS.

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for HTRA2/Omi Antibody (229926) [Unconjugated]

• High temperature requirement protein A2
• HtrA serine peptidase 2
• HTRA2
• HtrA-like serine protease
• Omi
• OMIOmi stress-regulated endoprotease
• PARK13EC 3.4.21.108
• PRSS25
• PRSS25serine, 25
• Serine protease 25
• serine protease HTRA2, mitochondrial
• Serine proteinase OMI

Background

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.

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.

Limitations

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.

Publications for HTRA2/Omi Antibody (MAB1458)(5)

Publications using MAB1458

• Taher Darreh-Shori, Sareh Rezaeianyazdi, Erica Lana, Sumonto Mitra, Anna Gellerbring, Azadeh Karami et al. Increased Active OMI/HTRA2 Serine Protease Displays a Positive Correlation with Cholinergic Alterations in the Alzheimer’s Disease Brain Molecular Neurobiology 7/1/2019 [PMID: 30361890]
• Taher Darreh-Shori, Sareh Rezaeianyazdi, Erica Lana, Sumonto Mitra, Anna Gellerbring, Azadeh Karami et al. Increased Active OMI/HTRA2 Serine Protease Displays a Positive Correlation with Cholinergic Alterations in the Alzheimer’s Disease Brain Molecular Neurobiology 2019-07-01 [PMID: 30361890] (ELISA Detection, Human)
• Serum HtrA1 is differentially regulated between early-onset and late-onset preeclampsia Placenta [PMID: 26187609] (ELISA Detection, N/A)
• Haudek SB, Taffet GE, Schneider MD, Mann DL TNF provokes cardiomyocyte apoptosis and cardiac remodeling through activation of multiple cell death pathways. J. Clin. Invest., 2007-09-01;117(9):2692-701. 2007-09-01 [PMID: 17694177] (Western Blot, Mouse)
• Setoguchi K, Otera H, Mihara K Cytosolic factor- and TOM-independent import of C-tail-anchored mitochondrial outer membrane proteins. EMBO J., 2006-11-16;25(24):5635-47. 2006-11-16 [PMID: 17110923] (ICC, Western Blot, Human)

Bioinformatics

• Gene Symbol: HTRA2
• Uniprot:
  • O43464()