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H6908

Manufactured by Merck Group
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Sourced in United Kingdom, United States, Germany

H6908 is a laboratory equipment product manufactured by Merck Group. It is a multipurpose device designed for use in various scientific applications. The core function of H6908 is to facilitate controlled and precise measurements or manipulations within a laboratory setting.

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Lab products found in correlation

F7425, by Merck Group (11 mentions) F1804, by Merck Group (9 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (8 mentions) H3663, by Merck Group (5 mentions) Anti ha, by Merck Group (4 mentions) Ab8227, by Abcam (4 mentions) Ab1791, by Abcam (4 mentions) Alexa fluor 488, by Thermo Fisher Scientific (4 mentions) Rabbit anti ha antibody, by Merck Group (3 mentions) A1978, by Merck Group (3 mentions) A2095, by Merck Group (3 mentions) A11034, by Thermo Fisher Scientific (3 mentions) Lsm 710 confocal microscope, by Zeiss (3 mentions) Hoechst 33342, by Thermo Fisher Scientific (3 mentions) Anti β actin, by ABclonal (2 mentions) Supersignal west femto maximum sensitivity substrate, by Thermo Fisher Scientific (2 mentions) Nitrocellulose membrane, by Bio-Rad (2 mentions) Passive lysis buffer (plb), by Promega (2 mentions) Hybond ecl nitrocellulose membrane, by GE Healthcare (2 mentions) M4439, by Merck Group (2 mentions)

82 protocols using h6908

1

Western Blot and Immunofluorescence Assays

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Total protein were extracted using RIPA lysis buffer (Beyotime, China). Proteins were separated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE), and transferred onto PVDF membranes (Millipore, Billerica, MA, USA). The membranes were then incubated with primary antibodies against the HA tag (1:2000, H6908, Sigma-Aldrich, Munich, Germany), Flag tag (1:2000, F2555, Sigma), METTL3 (1:1000, ab195352, Abcam, Cambridge, MA, USA), or GAPDH (1:1000, ab8245, Abcam) at 4 °C overnight. After incubation with species-matched secondary antibodies, the immunoreactive proteins were detected using chemiluminescence in a Gel imaging system (Bio-Rad, ChemiDoc MP Imaging System). For immunofluorescence, cells overexpressing ZNF750 were seeded and cultured on cover glass, and then fixed by methanol after 24 h of culture. The cells were then reacted with anti-HA antibody (1:500, H6908, Sigma), followed with the corresponding secondary fluorescent antibody. The cells were viewed using confocal microscopy (Olympus, Tokyo, Japan).
Zhang P., He Q., Lei Y., Li Y., Wen X., Hong M., Zhang J., Ren X., Wang Y., Yang X., He Q., Ma J, & Liu N. (2018). m6A-mediated ZNF750 repression facilitates nasopharyngeal carcinoma progression. Cell Death & Disease, 9(12), 1169.
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2

Immunofluorescence and Western Blotting Protocols

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Antibodies used for immunofluorescence in brain slices include anti-Nesprin-2 (1:1000, gift from Dr. Gundersen, [23 (link)]), anti-HA (Sigma-Aldrich, H6908, 1:2000) and donkey fluorophore-conjugated secondary antibodies (Jackson Labs, 1:500).
Antibodies used for immunofluorescence in cells include anti-dynein intermediate chain clone 74.1 (University of Virginia, 1:250), anti-BicD1 (1:250, Abcam, ab170878), anti-BicD2 (1:250, Abcam, ab117818), anti-Nde1/Ndel1 (1:250, [59 (link)]), anti-cyclin B1 (1:100, BD Biosciences, 554177), chicken polyclonal against GFP (1:150, Millipore, 16901), and donkey fluorophore-conjugated secondary antibodies (Jackson Labs, 1:250 dilution).
Antibodies used for western blotting include anti GST (1:3000, Santa Cruz, sc-53909), anti GFP (1:3000, Invitrogen, A11122), anti-BicD2 (1:2000, Abcam, ab117818), anti BicD2 CT (1:2000, GeneTex, GTX120683). To develop in a LI-COR system, fluorescent secondary antibodies (1:10,000) were acquired from Invitrogen and Rockland.
Gonçalves J.C., Quintremil S., Yi J, & Vallee R.B. (2020). Nesprin-2 Recruitment of BicD2 to the Nuclear Envelope Controls Dynein/Kinesin-mediated Neuronal Migration in Vivo. Current biology : CB, 30(16), 3116-3129.e4.
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3

Antibody Characterization and Signaling Pathway Analysis

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Mouse monoclonal antibodies against Flag (Origene, 1:2000, F3165), HA (Origene, 1:2000, H6908), β-actin (Sigma, 1:10,000, A2228), Myc (CST, 1:1000, 5605), p-IκBα (CST, 1:1000, 9246 L); rabbit antibodies against p-IRF3 (CST, 1:500, 37829), USP19 (Abcam, 1:1000, ab93159), TRIF (Abcam, 1:1000, ab180689), p65 (Santa Cruz Biotechnology, 1:1000, 71675), p-p65(S536) (CST, 1:1000, 3033),TBK1 (Abcam, 1:1000, ab40676) and p-TBK1 (Abcam, 1:1000, ab109272), ubiquitin (Abcam, 1:500, ab134953), K27-linkage specific polyubiquitin (Abcam, 1:1000, 181537), TLR3 (CST, 1:500, 6961), TLR4 (R&D, 1:500, AF1478), TRAM (Abcam, 1:500, ab96106), KCTD10 (Proteintech, 1:1000, 27279–1-AP); poly(I:C) (Invivogen), LPS (Sigma), R848 (Invivogen), PGN (Invivogen), human IFN-γ (Peprotech), murine M-CSF (Peprotech), Trizol (Takara Bio), SYBR Green (BIO-RAD), dual-specific luciferase assay kit (Promega, E1980), polybrene (Millipore,TR-1003-G), type II collagenase (Worthington), DNase I (Sigma-Aldrich), and d-galactosamine hydrochloride (D-Gal) (Sigma); and ELISA kits for TNF (Biolegend), IL-6 (Biolegend), CXCL10 (Boster) and IFN-β (PBL) were purchased from the indicated companies. HEK293 cells were obtained from ATCC. SeV (Cantell strain) (Charles River Laboratories), HSV-1 (KOS strain) (China Center for Type Culture Collection, Wuhan, China) were obtained from the indicated companies.
Wu X., Lei C., Xia T., Zhong X., Yang Q, & Shu H.B. (2019). Regulation of TRIF-mediated innate immune response by K27-linked polyubiquitination and deubiquitination. Nature Communications, 10, 4115.
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4

Antibody Panel for Protein Analysis

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Antibodies used in the study were as follows: anti-Flag (F3165/F7425, Sigma), anti-Myc (9B11/71D10, Cell Signaling Technology (CST)), anti-HA (16B12, Biolegend and H6908, Sigma), anti-TRAF2 (sc-136999, Santa Cruz), anti-Rab5 (sc-46692, Santa Cruz), anti-Rab7 (sc-376362, Santa Cruz), anti-Sprouty 2 (sc-100862, Santa Cruz/ab85670, Abcam), anti-EGFR (sc-373746, Santa Cruz), anti-p-Tyr (9411, CST), anti-p-Ser/Thr (ab9344, Abcam and 05-368, Sigma), anti-Ser (sc-81514, Santa Cruz), anti-Thr (sc-5267, Santa Cruz), anti-Ubiquitin (sc-8017, Santa Cruz), anti-β-Actin (AC026, Abclonal), anti-GFP (AE011, Abclonal), anti-glutathione S-transferase (GST) (2622 S, CST), anti-V5 (R960-25, Thermo Fisher and 30801ES10, Yeasen), anti-Mouse/Rabbit IgG-peroxidase secondary antibody (A0545/A9044, Sigma), anti-Mouse IgG (light chain specific)-peroxidase secondary antibody (115-005-174, Jackson), and anti-DYRK1A polyclonal antibody has been previously described [30 (link)].
Zhang P., Zhang Z., Fu Y., Zhang Y., Washburn M.P., Florens L., Wu M., Huang C., Hou Z, & Mohan M. (2021). K63-linked ubiquitination of DYRK1A by TRAF2 alleviates Sprouty 2-mediated degradation of EGFR. Cell Death & Disease, 12(6), 608.
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5

Western Blot Analysis of Mitochondrial Proteins

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Laemmli buffer containing 4% SDS, 10% β-mercaptoethanol (M6250; Sigma-Aldrich), 20% glycerol (GI345; Melford), 0.004% blue bromophenol (A2331,0025; AppliChem), and 0.125 M Tris-HCl was added to protein lysates, followed by boiling at 95°C for 10 min. Samples were separated on SDS-PAGE and transferred to a nitrocellulose (Macherey-Nagel) or a polyvinylidene difluoride (Merck) membrane. The membranes were blocked with 5% milk (A0830; PanReac AppliChem) and probed with the following antibodies: anti–β-Actin (ab8227; Abcam), anti–β-Tubulin (ab15568; Abcam), anti-CBS (14787-1-AP; Proteintech), anti-CSE (12217-1-AP; Proteintech), anti-MPST (HPA001240; Atlas Antibodies), anti-MTCO1 (ab14705; Abcam), anti-Citrate Synthase (ab96600; Abcam), anti-SOD2 (13141; Cell Signaling), anti-TOM40L (ab236421; Abcam), anti-TIM50 (ab109436; Abcam), anti-HIF1α (sc-13515; Santa Cruz Biotechnology), anti-H3 (ab176842; Abcam), anti-V5 (Merck), anti-HA (H6908; Sigma-Aldrich), and anti-Biotin (HRP conjugate; 5571; Cell Signaling). Immunoblots were next incubated with a secondary antibody (anti-rabbit [AP132P; Merck] or anti-mouse [7076; Cell Signaling]) and visualized using the Western HRP substrate (Merck). ImageJ software was used to quantify the expression levels of the proteins.
Katsouda A., Valakos D., Dionellis V.S., Bibli S.I., Akoumianakis I., Karaliota S., Zuhra K., Fleming I., Nagahara N., Havaki S., Gorgoulis V.G., Thanos D., Antoniades C., Szabo C, & Papapetropoulos A. (2022). MPST sulfurtransferase maintains mitochondrial protein import and cellular bioenergetics to attenuate obesity. The Journal of Experimental Medicine, 219(7), e20211894.
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6

Western Blot Analysis of IFI6 Protein

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Cells were lysed in passive lysis buffer (Promega) and clarified by centrifugation. Cell lysates or recombinant IFI6 protein were mixed with Laemmli sample buffer containing 2.5% β-mercaptoethanol, and heated at 95°C for 5 min, before SDS-PAGE electrophoresis. Proteins were transferred to nitrocellulose membranes (Biorad), and detected using primary rabbit polyclonal antibodies (pAbs) specific for the HA epitope tag (Sigma Aldrich H6908), GST epitope tag (Sigma Aldrich A7340), IFI6 (ABclonal A6157), IFI6 (St John´s laboratory STJ27910 and STJ 191871) and mouse monoclonal antibodies (mAbs) against the FLAG epitope tag (Sigma-Aldrich F3165), His epitope tag (ThermoFisher Scientific MA1-21315), GFP (Merck 11814460001), actin (Sigma-Aldrich A1978), and ubiquitin (Santa Cruz Biotechnology, sc-166553); following by binding to goat anti-rabbit (pAb) or anti-mouse (mAb) IgG antibodies (Abs) conjugated to horseradish peroxidase (Sigma-Aldrich), at a 1:4,000 dilution. nitrocellulose membranes were revealed by chemiluminescence with the SuperSignal west femto maximum sensitivity substrate (ThermoFisher Scientific), according to the manufacturer’s recommendations. Where indicated, protein bands have been quantified by densitometry using the ImageJ (Fiji) software.
Villamayor L., Rivero V., López-García D., Topham D.J., Martínez-Sobrido L., Nogales A, & DeDiego M.L. (2023). Interferon alpha inducible protein 6 is a negative regulator of innate immune responses by modulating RIG-I activation. Frontiers in Immunology, 14, 1105309.
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7

Immunoprecipitation and Fractionation Protocol

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For immunoprecipitation, cells were lysed on ice in TNE buffer (50 mM Tris–HCl, pH 7.5, 150 mM NaCl, 0.5% Tergitol-type NP-40, 1 mM EDTA, and protease inhibitors). Anti-FLAG M2 magnetic beads (M8823, Sigma) were incubated overnight with cell lysate fractions. Samples were then washed 6 times with TBS. Western blotting was performed following standard principles and immunofluorescence was assessed using a Leica TCS SP2 spectral confocal microscope. Nuclear-cytoplasmic fractionation was performed following standard protocol. The following primary antibodies were used: anti-FLAG (F7425, Sigma), anti-NCoR (ABE251, Millipore), anti-HA (H6908, Sigma), anti-SMRT (ab24551, Abcam), anti-HDAC7 (ab12174, Abcam), anti-E-cadherin (BD Biosciences), anti-SSEA-1 (MC480, Cell Signaling), anti-SOX2 (MAB2018, R&D Systems), anti-OCT4 (sc-8628, Santa Cruz), anti-KLF4 (AF3158, R&D Systems), anti-MYC (AF3696, R&D Systems), anti-histone H3 (ab1791, Abcam), anti-ACTIN (A5316, Sigma), and anti-GAPDH (G8795, Sigma). DAPI was purchased from Sigma (D9542).
Luo Z., Qing X., Benda C., Huang Z., Zhang M., Huang Y., Zhang H., Wang L., Lai Y., Ward C., Volpe G., Zhong X., Qin B., Zhuang Q., Esteban M.A, & Li W. (2019). Nuclear-cytoplasmic shuttling of class IIa histone deacetylases regulates somatic cell reprogramming. Cell Regeneration, 8(1), 21-29.
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8

HA-Tagged Protein Detection in C. reinhardtii

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For Western blot analysis, protein samples of mid-log phase C. reinhardtii at equal cell density were prepared and separated by SDS-PAGE using a gel containing 15% acrylamide as described in Young and Purton (2014 (link)). The proteins were blotted onto a Hybond ECL nitrocellulose membrane (GE Healthcare) using a Trans-Blot SD semi-dry electrophoretic transfer cell (Bio-Rad) at 19 V for 1 h. The membrane was blocked overnight in TBS-T (TBS + 0.1% Tween) with 5% milk and incubated with a primary antibody (α-HA antibody produced in rabbit diluted at 1:2000, Sigma-Aldrich product H6908) for 1 h. After washing the membrane in TBS-T for 30 min (5–15 min each time), it was incubated with a secondary antibody (Goat anti-rabbit IgG, DyLight 800 diluted at 1:25,000, Thermo Scientific product 35571) for 1 h and followed again by washing in TBS-T. Both antibodies were diluted in TBS-T with 0.5% milk. For detection, the membrane was analysed using the Odyssey Infrared Imaging system (Li-COR Biosciences).
Changko S., Rajakumar P.D., Young R.E, & Purton S. (2019). The phosphite oxidoreductase gene, ptxD as a bio-contained chloroplast marker and crop-protection tool for algal biotechnology using Chlamydomonas. Applied Microbiology and Biotechnology, 104(2), 675-686.
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9

Visualizing MEF2C and UBE3A Localization

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Expression plasmids expressing human MEF2C and UBE3A and respective negative control plasmids were obtained from Sino Biologicals (MEF2C-HA: HG12320-CY, UBE3A-Myc:HG11130-CM, pCMV-Myc:CV014 and pCMV-HA:CV013) and used for transient transfection. Transfected HeLa cells were grown on poly-lysine coated coverslips, fixated with 4% paraformaldehyde in PBS for 10 minutes and stained with anti-Myc (M4439, Sigma-Aldrich) and anti-HA (H6908, Sigma-Aldrich) and with Alexa Fluor™ 488 goat anti–mouse and Alexa Fluor™ 488 donkey anti–rabbit antibodies (A11001 and A10040, Thermo Fisher). Nuclei were counterstained with DAPI (Serva). Images were taken with a Zeiss Axio Imager Z2 Apotome microscope with a 63x objective and analyzed in ImageJ.
Straub J., Gregor A., Sauerer T., Fliedner A., Distel L., Suchy C., Ekici A.B., Ferrazzi F, & Zweier C. (2020). Genetic interaction screen for severe neurodevelopmental disorders reveals a functional link between Ube3a and Mef2 in Drosophila melanogaster. Scientific Reports, 10, 1204.
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10

JMJD6 Protein Interactions in HEK293T Cells

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HEK293T cells were transfected at a confluency of 60% with C-terminal HA-tagged JMJD6-2, JMJD6-Ex5 or JMJD6-2-ΔpolyS and GFP-tagged U2AF65, U2AF6520-70 or UBF (26672, Addgene, Watertown, MA, USA) using Lipofectamine 2000 (11668030, Thermo Fisher, Waltham, MA, USA). The same experiment was conducted with HEK293T cells transfected with GFP-tagged JMJD6-2, JMJD6-Ex5 or JMJD6-2-ΔpolyS and Flag-tagged FCP1. Beads with anti-GFP (ABIN509397, ChromoTek, Planegg-Martinsried, Germany) were used for immunoprecipitation as described in Webby et al. (2009) [6 (link)]. Western blots were stained with mouse anti-GFP antibody (11814460001, Roche, Basel, Switzerland), rabbit anti-HA antibody (H6908, Sigma Aldrich, St. Louis, MO, USA) and mouse anti-Flag antibody (F1804, Sigma Aldrich, St. Louis, MO, USA).
Raguz N., Heim A., Engal E., Wesche J., Merl-Pham J., Hauck S.M., Erkelenz S., Schaal H., Bensaude O., Wolf A., Salton M, & Böttger A. (2020). JMJD6 Regulates Splicing of Its Own Gene Resulting in Alternatively Spliced Isoforms with Different Nuclear Targets. International Journal of Molecular Sciences, 21(18), 6618.
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