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Ab151769

Manufactured by Abcam
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Ab151769 is a laboratory reagent used for research purposes. It is a monoclonal antibody that targets a specific protein or antigen. The core function of this product is to facilitate the detection and analysis of the target molecule in various experimental settings.

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Lab products found in correlation

Passive lysis buffer (plb), by Promega (5 mentions) Pvdf membrane, by Cytiva (4 mentions) Ab135626, by Abcam (4 mentions) Ab1376, by Abcam (2 mentions) Ab140600, by Abcam (2 mentions) Actin, by Cell Signaling Technology (2 mentions) P stat5, by Cell Signaling Technology (2 mentions) β tubulin 9f3, by Cell Signaling Technology (2 mentions) Stat5, by Santa Cruz Biotechnology (2 mentions) Creb 2 c 20, by Santa Cruz Biotechnology (2 mentions) Hrp conjugated anti rabbit secondary antibody, by Agilent Technologies (2 mentions) Ab85377, by Abcam (2 mentions) Enhanced chemi luminescence (ecl), by Merck Group (1 mentions) Ab8245, by Abcam (1 mentions) P0013, by Beyotime (1 mentions) Polyvinylidene difluoride membrane, by Cytiva (1 mentions) Horseradish peroxidase conjugated anti rabbit secondary antibody, by Abcam (1 mentions) Pageruler plus prestained, by Thermo Fisher Scientific (1 mentions) Amersham imager 600, by GE Healthcare (1 mentions) Tubulin, by Cell Signaling Technology (1 mentions)

13 protocols using ab151769

1

Quantification of Protein Levels

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Total proteins were obtained from the cells or tissues on ice using a lysis buffer (P0013, Beyotime Biotechnology), separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and then transferred to a polyvinylidene difluoride membrane. After blocking the membrane and treating with mammalian target of CSE (ab151769), CBS (ab140600), nNOS (ab1376), or GAPDH (ab8245, 1 : 1000, Abcam) antibodies for 3 h at room temperature, the membrane was incubated with the secondary antibody (1 : 3000, Abcam). Finally, the protein expression was evaluated using enhanced chemiluminescence (ECL, Millipore, USA).
Liu Y., Wei W., Liang S., Fang H, & Cao J. (2022). Esculentoside A Alleviates Intestinal Dysmotility in Ulcerative Colitis by Regulating H2S/CSE and NO/nNOS Systems. Evidence-based Complementary and Alternative Medicine : eCAM, 2022, 7757833.
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2

Immunohistochemical Analysis of CBS, CSE, and nNOS

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The sections were acquired similarly as in the H&E staining procedures. After dewaxing and hydration, the slides were exposed to citrate buffer. The slides were then pretreated at 80°C and incubated with 3% H2O2 to block endogenous peroxide. Thereafter, the slides were treated with anti-cystathionine β-synthase (CBS, ab140600, Abcam) antibody, anti-cystathionine γ-lyase (CSE, ab151769, Abcam) antibody, or antineuronal nitric oxide synthase (nNOS, ab1376, Abcam) antibody for 2 h at room temperature. After incubation with the relevant secondary antibody, the slides were mounted and analyzed using a light microscope (Nikon).
Liu Y., Wei W., Liang S., Fang H, & Cao J. (2022). Esculentoside A Alleviates Intestinal Dysmotility in Ulcerative Colitis by Regulating H2S/CSE and NO/nNOS Systems. Evidence-based Complementary and Alternative Medicine : eCAM, 2022, 7757833.
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3

Western Blot Analysis of Sulfur Metabolism

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Protein analysis was performed via western blotting on tissue homogenates in passive lysis buffer (Promega), separated by SDS-polyacrylamide gel electrophoresis, transferred to polyvinylidene difluoride membrane (Whatman) and blotted for CGL (ab151769 Abcam), CBS (ab135626 Abcam), MST (#85211 Abcam), or α-Tubulin (#4074 Abcam), followed by horseradish peroxidase-conjugated secondary anti-rabbit antibody (#97051 Abcam). Proteins were visualized using SuperSignal West Femto Maximum Sensitivty Substrate (Thermo Scientific #34096) on an Amersham Imager 600 (General Electric) and sizes determined using the PageRuler Plus Prestained (26619 Thermo Fisher). After blotting and washing, the membranes were stained using Coomassie blue, to determine the relatively equal amount of tissue proteins loaded.
Yang J., Minkler P., Grove D., Wang R., Willard B., Dweik R, & Hine C. (2019). Non-enzymatic hydrogen sulfide production from cysteine in blood is catalyzed by iron and vitamin B6. Communications Biology, 2, 194.
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4

Western Blot Analysis of Cellular Stress Markers

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Cells were homogenized with passive lysis buffer (Promega), normalized for protein content, boiled with SDS loading buffer and separated by SDS-PAGE. Proteins were transferred to PVDF membrane (Whatman) and blotted for CGL (ab151769 Abcam), HIF1α (10006421 Cayman Chemical), p-eIF2α Ser51 (9712S Cell Signaling), total eIF2α(9722S Cell Signaling), ATF4 (11815 Cell Signaling), Actin (13E5 Cell Signaling) and Tubulin (2146S Cell Signaling) and secondarily with HRP-conjugated anti-rabbit antibody (Dako).
Longchamp A., Mirabella T., Arduini A., MacArthur M.R., Das A., Treviño-Villarreal J.H., Hine C., Ben-Sahra I., Knudsen N.H., Brace L.E., Reynolds J., Mejia P., Tao M., Sharma G., Wang R., Corpataux J.M., Haefliger J.A., Ahn K.H., Lee C.H., Manning B.D., Sinclair D.A., Chen C.S., Ozaki C.K, & Mitchell J.R. (2018). Amino acid restriction triggers angiogenesis via GCN2/ATF4 regulation of VEGF and H2S production. Cell, 173(1), 117-129.e14.
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5

Comprehensive Protein Expression Analysis

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Protein analysis was performed via Western blot on tissue and cell homogenates in passive lysis buffer (Promega), separated by SDS-PAGE, transferred to PVDF membrane (Whatman) and blotted for CGL (ab151769 Abcam), CBS (ab135626 Abcam), 3MST (HPA001240 Sigma), Stat5 (sc-835 Santa Cruz), p-Stat5 (#9359 Cell Signaling Technology), GNMT (Aviva), AHCY (Abcam ab56146), ATF4 (aka CREB-2 C-20, Santa Cruz Biotechnology sc-200), ATG5 (Novus, NB110–53818), ATG7 (Sigma, A2856), β-Tubulin 9F3 (#2128 Cell Signaling) or Actin (#4970 Cell Signaling) followed by HRP conjugated secondary anti-rabbit antibody (Dako).
Hine C., Kim H.J., Zhu Y., Harputlugil E., Longchamp A., Matos M.S., Ramadoss P., Bauerle K., Brace L., Asara J.M., Ozaki C.K., Cheng S.Y., Singha S., Ahn K.H., Kimmelman A., Fisher F.M., Pissios P., Withers D.J., Selman C., Wang R., Yen K., Longo V.D., Cohen P., Bartke A., Kopchick J.J., Miller R., Hollenberg A.N, & Mitchell J.R. (2017). Hypothalamic-Pituitary Axis Regulates Hydrogen Sulfide Production. Cell Metabolism, 25(6), 1320-1333.e5.
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6

Comprehensive Protein Analysis Protocol

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Protein analysis was performed via western blot on tissue and cell homogenates in passive lysis buffer (Promega), separated by SDS-PAGE, transferred to PVDF membrane (Whatman) and blotted for CGL (ab151769 Abcam), CBS (ab135626 Abcam), 3MST (HPA001240 Sigma), Stat5 (sc-835 Santa Cruz), p-Stat5 (#9359 Cell Signaling Technology), GNMT (Aviva), AHCY (Abcam ab56146), ATF4 (aka CREB-2 C-20, Santa Cruz Biotechnology sc-200), ATG5 (Novus, NB110-53818), ATG7 (Sigma, A2856), β-Tubulin 9F3 (#2128 Cell Signaling) or Actin (#4970 Cell Signaling) followed by HRP conjugated secondary anti-rabbit antibody (Dako).
Hine C., Kim H.J., Zhu Y., Harputlugil E., Longchamp A., Matos M.S., Ramadoss P., Bauerle K., Brace L., Asara J.M., Ozaki C.K., Cheng S.Y., Singha S., Ahn K.H., Kimmelman A., Fisher F.M., Pissios P., Withers D.J., Selman C., Wang R., Yen K., Longo V.D., Cohen P., Bartke A., Kopchick J.J., Miller R., Hollenberg A.N, & Mitchell J.R. (2017). Hypothalamic-Pituitary Axis Regulates Hydrogen Sulfide Production. Cell Metabolism, 25(6), 1320-1333.e5.
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7

Kidney Ischemia-Reperfusion Model in Rats

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To develop the kidney I/R model, male Sprague Dawley (S-D) rats at 10–12 weeks of age were used. Male Wistar Kyoto (WKY) and spontaneously hypertensive rats (SHRs) aged 16–18 weeks were used for detecting the role of NW in hypertension. All of rats were kept in a temperature- and humidity-controlled room and had free access to tap water. All animal protocols complied with all relevant ethical regulations and were approved by the Institutional Animal Care and Use Committee, Experimental Animal Center, Fuwai Hospital, National Center for Cardiovascular Diseases, China. DMEM, DMEM/F12 culture medium, and TRIZOL reagent were from Invitrogen. Primary antibodies for β-actin (SC-47778) and Eif5 (SC-28309) were from Santa Cruz Biotechnology. The anti-CSE antibody (ab151769) was from Abcam. DL-Propargylglycine (PPG) (P7888) was from Sigma-Aldrich. NW (54954-12-0) was from Lookchem Biological Technology Co., Ltd.
Niu Y., Du C., Cui C., Zhang H., Deng Y., Cai J., Chen Z, & Geng B. (2021). Norswertianolin Promotes Cystathionine γ-Lyase Activity and Attenuates Renal Ischemia/Reperfusion Injury and Hypertension. Frontiers in Pharmacology, 12, 677212.
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8

CRISPR-Cas9 Knockout of CSE, SQR, and MST

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The following guide RNAs (gRNAs) were used in the study: For CSE: exon 5 (5’-GGAGACATTATTTTGGTCGTGG-3’) and exon 1 (5’-TTCCAACATTTGGCCACGCAGG-3’) (purchased from Sigma-Aldrich); for SQR: exon 8 (5’-GTCCTCAAGACCAGTCCTGTGG-3’) (Sigma-Aldrich) and exon 3 (5’-TGCCGTGGGACGACCAGATG-3’) (Genscript); for MST: exon 4 (5’-CGCGTTACCGTCTCGGGGCT-3’) (Genscript). The gRNAs were expressed using an U6gRNA-Cas9-2A-RFP (Sigma-Aldrich), or pSpCas9 BB-2A-Puro (PX459) v2.0 vector after transient transfection using Lipofectamine LTX following the manufacturer’s protocol. Depending on the vector, cells were selected either with 12.5 μg/ml puromycin (Sigma), or by FACS. Individual clones were grown after diluting cells to a concentration of 0.5 cells per 100 μl in 96 well plates. Knockout clones were identified by western blotting. Antibodies used were rabbit anti-CSE (abcam, ab151769) at a dilution of 1:500, mouse anti-SQR (abcam, ab71978) at a dilution of 1:500 and mouse anti-MST (Santa Cruz, sc-374326) at a dilution of 1:100. Rabbit anti-GAPDH (abcam, ab 9485) (1:10000) and mouse anti-β-actin (sigma # A5441) (1:2500) were used as loading controls.
Ezeriņa D., Takano Y., Hanaoka K., Urano Y, & Dick T.P. (2018). N-acetyl cysteine functions as a fast-acting antioxidant by triggering intracellular H2S and sulfane sulfur production. Cell chemical biology, 25(4), 447-459.e4.
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9

Western Blot Analysis of Neurotransmitter Transporters

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All chemicals were purchased from Sigma-Aldrich (St Louis, MO, USA) unless otherwise indicated. Methylarsonic acid (MMAV) disodium salt (99% pure) was obtained from Chem Service (West Chester, PA, USA). Sodium borohydride was obtained from EM Science (Gibbstown, NJ, USA). For Western blots, primary rabbit antibodies against xCT (ab37185), EAAT3 (ab124802), GLAST (ab416), GLT1 (ab41621), NR2B (ab65783), GluA2 (ab133477), Synaptophysin (SY38, ab8049), CBS (ab135626), and CSE (ab151769) were obtained from Abcam (Cambridge, MA, USA). Anti-LAT1 (sc-134994) and PSD95 (7E3, sc-32290) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Rabbit anti-NR2A (AB1555P) and anti-GluA1 (AB1504) antibodies were purchased from Millipore (Bedford, MA, USA). Primary antibody against SLC1A4 (8442 s) and secondary goat anti-rabbit antibodies were obtained from Cell Signaling Technology (Danvers, MA, USA). Secondary anti-mouse IgG antibody was purchased from Invitrogen (Waltham, MA, USA).
González-Alfonso W.L., Pavel P., Karina H.M., Del Razo L.M., Sanchez-Peña L.C., Zepeda A, & Gonsebatt M.E. (2023). Chronic exposure to inorganic arsenic and fluoride induces redox imbalance, inhibits the transsulfuration pathway, and alters glutamate receptor expression in the brain, resulting in memory impairment in adult male mouse offspring. Archives of Toxicology, 97(9), 2371-2383.
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10

Quantifying Sulfur Metabolism Proteins

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Total RNA was isolated from tissues and cells using miRNeasy Mini Kit (Qiagen) and cDNA synthesized by random hexamer priming with the Verso cDNA kit (Thermo). qRT-PCR was performed with SYBR green dye (Lonza) and TaqPro DNA polymerase (Denville). Fold changes were calculated by the ΔΔCt method using Hprt and/or Rpl13 as standards and normalized to the experimental WT AL control. Protein expression was analyzed in tissues homogenized in passive lysis buffer (Promega), separated by SDS-PAGE, transferred to PVDF membrane (Whatman) and blotted for CGL (ab151769 Abcam), CBS (ab135626 Abcam) or Actin (13E5 Cell Signaling) followed by HPRT conjugated secondary anti-rabbit antibody (Dako).
Hine C., Harputlugil E., Zhang Y., Ruckenstuhl C., Lee B.C., Brace L., Longchamp A., Trevino-Villarreal J.H., Mejia P., Ozaki C.K., Wang R., Gladyshev V.N., Madeo F., Mair W.B, & Mitchell J.R. (2014). Endogenous Hydrogen Sulfide Production Is Essential for Dietary Restriction Benefits. Cell, 160(0), 132-144.
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