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Protein block

Manufactured by Thermo Fisher Scientific
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The Protein block is a laboratory equipment designed for the separation and analysis of proteins. Its core function is to provide a stable and controlled environment for the electrophoretic separation of proteins based on their molecular weight or charge. The device allows for the effective resolution and visualization of protein samples, enabling researchers to study and characterize these biomolecules.

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Lab products found in correlation

Target retrieval buffer, by Vector Laboratories (5 mentions) Peroxidase blocking buffer, by Thermo Fisher Scientific (5 mentions) Biotinylated anti rabbit or polyvalent secondary antibody, by Thermo Fisher Scientific (5 mentions) S0809, by Agilent Technologies (4 mentions) Mouse on mouse m o m blocking reagent mkb 2213, by Vector Laboratories (2 mentions) Biotinylated secondary antibody, by Thermo Fisher Scientific (1 mentions) H 3300, by Vector Laboratories (1 mentions) Mayer s hematoxylin, by Merck Group (1 mentions) Hydrogen peroxide block, by Thermo Fisher Scientific (1 mentions) Gdf 15, by Merck Group (1 mentions) Anti p53 do 7, by Agilent Technologies (1 mentions) Organosilane coated slides, by Merck Group (1 mentions) Anti tnf α, by Santa Cruz Biotechnology (1 mentions) Permount, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

I-Block Protein-Based Blocking Reagent Serum-free protein block UltraVision Protein Block Ultravision Protein Block solution Hydrogen Peroxide Block and Protein Block

11 protocols using protein block

1

Immunohistochemistry Staining Protocol

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All antibodies are described in the Supplementary Information. Patient samples were collected under IRB exemption approval for protocol #EX21205258-1. Paraffin embedded sections were rehydrated through a xylene and alcohol series, rinsed in H2O and washed in PBS. Antigen retrieval was performed using target retrieval buffer (Vector Labs, Burlingame, CA) and steamed for 20 min. Samples were then blocked in a peroxidase blocking buffer (Thermo Scientific) for 15 min, followed by Protein block (Thermo Scientific) for 5 min, and incubated in appropriate primary antibody diluted in antibody diluent (S0809, Dako) at 4°C overnight in a humidified chamber. For mouse samples to be incubated with anti-mouse antibody, samples were blocked for 1 hour in Mouse on Mouse (M.O.M.™) Blocking Reagent (MKB-2213, Vector Labs). Following washing in PBS, samples were incubated in biotinylated anti-rabbit or polyvalent secondary antibody (Thermo Scientific) followed by streptavidin-HRP solution at room temperature for 20 min. Samples were then washed in PBS and incubated in 3-Amino-9-Ethyl-l-Carboazole (AEC) chromogen and counter stained with Mayers hematoxylin for 1 min, rinsed in cold H2O, and mounted in Aquamount.
Kaur A., Webster M.R., Marchbank K., Behera R., Ndoye A., Kugel CH I.I.I., Dang V.M., Appleton J., O’Connell M.P., Cheng P., Valiga A.A., Morissette R., McDonnell N.B., Ferrucci L., Kossenkov A.V., Meeth K., Tang H.Y., Yin X., Wood WH I.I.I., Lehrmann E., Becker K.G., Flaherty K.T., Frederick D.T., Wargo J.A., Cooper Z.A., Tetzlaff M.T., Hudgens C., Aird K.M., Zhang R., Xu X., Liu Q., Bartlett E., Karakousis G., Eroglu Z., Lo R.S., Chan M., Menzies A.M., Long G.V., Johnson D.B., Sosman J., Schilling B., Schadendorf D., Speicher D.W., Bosenberg M., Ribas A, & Weeraratna A.T. (2016). sFRP2 in the aged microenvironment drives melanoma metastasis and therapy resistance. Nature, 532(7598), 250-254.
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2

Immunofluorescence Staining of DUX4 in Cells

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Cells were fixed with 3.8% PFA, washed three times, permeabilised in 0.5% (v/v) Triton X-100 in PBS for 7 min, and washed with washing buffer (0.1% (v/v) Tween20 in PBS). The samples were incubated with ProteinBlock (Thermo Fisher Scientific) at room temperature for 10 min to prevent unspecific binding of primary antibody. Primary antibody (rabbit MAb anti-DUX4, clone E5-5, Abcam) was diluted 1:300 in washing buffer and incubated at 4°C overnight. After washings, fluorescence-conjugated secondary antibody (anti rabbit 594, A-21207; Thermo Fisher Scientific) was diluted 1:1000 in washing buffer and incubated at room temperature for 20 min. Nuclei were counterstained with DAPI 1:1000 in washing buffer. The images were captured with an Evos FL Cell Imaging system using 10× and 20× Plan Achromatic objectives.
Vuoristo S., Bhagat S., Hydén-Granskog C., Yoshihara M., Gawriyski L., Jouhilahti E.M., Ranga V., Tamirat M., Huhtala M., Kirjanov I., Nykänen S., Krjutškov K., Damdimopoulos A., Weltner J., Hashimoto K., Recher G., Ezer S., Paluoja P., Paloviita P., Takegami Y., Kanemaru A., Lundin K., Airenne T.T., Otonkoski T., Tapanainen J.S., Kawaji H., Murakawa Y., Bürglin T.R., Varjosalo M., Johnson M.S., Tuuri T., Katayama S, & Kere J. (2022). DUX4 is a multifunctional factor priming human embryonic genome activation. iScience, 25(4), 104137.
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3

Paraffin-Embedded Immunohistochemistry Protocol

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Immunohistochemistry was performed on paraffin embedded sections as
described previously (18 ). Briefly,
Paraffin embedded sections were rehydrated through a xylene and alcohol series
rinsed in H2O and washed in PBS. Antigen retrieval was performed by steaming for
20 min in target retrieval buffer (Vector Labs, Burlingame, CA) and steamed for
20 min. Samples were blocked in a peroxidase blocking buffer (Thermo Scientific)
followed by Protein block (Thermo Scientific) and incubated in appropriate
primary antibody diluted in antibody diluent (S0809, Dako) at 4°C
overnight in a humidified chamber. Samples were then incubated in biotinylated
anti-rabbit or polyvalent secondary antibody (Thermo Scientific) followed by
streptavidin-HRP solution. Samples were then washed in PBS and incubated in
3-Amino-9-Ethyl-l-Carboazole (AEC) chromogen. Finally, samples were washed in
H2O, incubated in Meyer’s hematoxylin for 1 min, rinsed in cold H2O, and
mounted in Aquamount. Patient samples were collected under IRB exemption
approval for protocol #EX21205258-1.
Behera R., Kaur A., Webster M.R., Kim S., Ndoye A., Kugel CH I.I.I., Alicea G.M., Wang J., Ghosh K., Cheng P., Lisanti S., Marchbank K., Dang V., Levesque M., Dummer R., Xu X., Herlyn M., Aplin A.E., Roesch A., Caino C., Altieri D.C, & Weeraratna A.T. (2017). Inhibition of age-related therapy resistance in melanoma by rosiglitazone-mediated induction of Klotho. Clinical cancer research : an official journal of the American Association for Cancer Research, 23(12), 3181-3190.
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4

Quantitative Characterization of DUX4 in Early Embryonic Development

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For characterization and quantitation of the DUX4 protein diploid zygotes (n = 3) and embryos (2-cell, n = 3; 4-cell, n = 4; 8-cell, n = 2 were fixed in 3.8 % PFA at room temperature for 15 min, washed three times in washing buffer (0.1% Tween 20 in PBS), and permeabilised in 0.5% Triton X-100 in PBS at room temperature for 15 min and washed once. Unspecific primary antibody binding was blocked with ProteinBlock (Thermo Fisher Scientific) by incubation at room temperature for 10 min. Primary antibody (rabbit MAb anti-DUX4, clone E5-5, Abcam) was diluted 1:300 in washing buffer and incubated at 4°C overnight. After three washes, the embryos were incubated in the secondary antibody (anti-rabbit Alexa 488, A-21206; Thermo Fisher Scientific) diluted 1:500 in washing buffer (as above) at room temperature for 2 h. After three washes, nuclei were counterstained with DAPI 1:500 in washing buffer.
Vuoristo S., Bhagat S., Hydén-Granskog C., Yoshihara M., Gawriyski L., Jouhilahti E.M., Ranga V., Tamirat M., Huhtala M., Kirjanov I., Nykänen S., Krjutškov K., Damdimopoulos A., Weltner J., Hashimoto K., Recher G., Ezer S., Paluoja P., Paloviita P., Takegami Y., Kanemaru A., Lundin K., Airenne T.T., Otonkoski T., Tapanainen J.S., Kawaji H., Murakawa Y., Bürglin T.R., Varjosalo M., Johnson M.S., Tuuri T., Katayama S, & Kere J. (2022). DUX4 is a multifunctional factor priming human embryonic genome activation. iScience, 25(4), 104137.
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5

Immunohistochemical Staining of Tissue Sections

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Sections were deparaffinized using xylene followed by rehydration through series of alcohol washes and finally PBS. Heat-mediated antigen retrieval was performed using citrate-based retrieval buffer (Vector Labs, H-3300). Samples were blocked in peroxide blocking buffer (Thermo Fisher Scientific), followed by protein block (Thermo Fisher Scientific) and incubated in appropriate antibody (Ki67; Novus, NB600-1252, Wnt5A; R&D Systems, mab645) at 4°C overnight in a humidified chamber. Following day, samples were washed and incubated with a biotinylated secondary antibody (Thermo Fisher Scientific) followed by streptavidin-HRP incubation. Samples were then washed in PBS and incubated in 3-amino-9-ethyl-1-carboazole chromogen and counterstained with Mayer’s hematoxylin (Millipore Sigma), rinsed with water and mounted in aquamount.
Douglass S.M., Fane M.E., Sanseviero E., Ecker B.L., Kugel CH I.I.I., Behera R., Kumar V., Tcyganov E.N., Yin X., Liu Q., Chhabra Y., Alicea G.M., Kuruvilla R., Gabrilovich D.I, & Weeraratna A.T. (2020). Myeloid-Derived Suppressor Cells Are a Major Source of Wnt5A in the Melanoma Microenvironment and Depend on Wnt5A for Full Suppressive Activity. Cancer research, 81(3), 658-670.
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6

Immunohistochemical Analysis of β-catenin, Hex, and Ki-67

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Immunohistochemical evaluation of β-catenin, Hex, and Ki-67 was performed on paraffin-embedded tissues. After antigen retrieval by boiling with 10mM sodium citrate (pH 6.0) endogenous peroxidase was blocked with Hydrogen Peroxide Block and then endogenous immunoglobulins was blocked Protein Block (Thermo Fisher Scientific, USA) according to manufacturer's protocol. The slides were incubated with primary antibodies to β-catenin (1 : 100, Cell Marque, USA), Hex (1 : 200, Santa Cruz Biotechnology, USA), and Ki-67 (1 : 100, Cell Marque, CA, USA) overnight at 8 °C. The reaction was visualized with anti-mouse/antirabbit “UltraVision LP Detection System” reagent kit (Thermo Fisher Scientific, USA) according to manufacturer's recommendations. The sections were counterstained with Mayer's hematoxylin. After dehydration the sections were mounted in polymer media. Sections processed without primary antibodies were used as negative control for secondary antibodies. No nonspecific reactions were registered.
Yaglova N.V., Tsomartova D.A., Obernikhin S.S., Nazimova S.V., Ivanova M.Y., Chereshneva E.V., Yaglov V.V, & Lomanovskaya T.A. (2021). Transcription factors β-catenin and Hex in postnatal development of the rat adrenal cortex: implication in proliferation control. Heliyon, 7(1), e05932.
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7

Immunohistochemistry Staining Protocol

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All antibodies are described in the Supplementary Information. Patient samples were collected under IRB exemption approval for protocol #EX21205258-1. Paraffin embedded sections were rehydrated through a xylene and alcohol series, rinsed in H2O and washed in PBS. Antigen retrieval was performed using target retrieval buffer (Vector Labs, Burlingame, CA) and steamed for 20 min. Samples were then blocked in a peroxidase blocking buffer (Thermo Scientific) for 15 min, followed by Protein block (Thermo Scientific) for 5 min, and incubated in appropriate primary antibody diluted in antibody diluent (S0809, Dako) at 4°C overnight in a humidified chamber. For mouse samples to be incubated with anti-mouse antibody, samples were blocked for 1 hour in Mouse on Mouse (M.O.M.™) Blocking Reagent (MKB-2213, Vector Labs). Following washing in PBS, samples were incubated in biotinylated anti-rabbit or polyvalent secondary antibody (Thermo Scientific) followed by streptavidin-HRP solution at room temperature for 20 min. Samples were then washed in PBS and incubated in 3-Amino-9-Ethyl-l-Carboazole (AEC) chromogen and counter stained with Mayers hematoxylin for 1 min, rinsed in cold H2O, and mounted in Aquamount.
Kaur A., Webster M.R., Marchbank K., Behera R., Ndoye A., Kugel CH I.I.I., Dang V.M., Appleton J., O’Connell M.P., Cheng P., Valiga A.A., Morissette R., McDonnell N.B., Ferrucci L., Kossenkov A.V., Meeth K., Tang H.Y., Yin X., Wood WH I.I.I., Lehrmann E., Becker K.G., Flaherty K.T., Frederick D.T., Wargo J.A., Cooper Z.A., Tetzlaff M.T., Hudgens C., Aird K.M., Zhang R., Xu X., Liu Q., Bartlett E., Karakousis G., Eroglu Z., Lo R.S., Chan M., Menzies A.M., Long G.V., Johnson D.B., Sosman J., Schilling B., Schadendorf D., Speicher D.W., Bosenberg M., Ribas A, & Weeraratna A.T. (2016). sFRP2 in the aged microenvironment drives melanoma metastasis and therapy resistance. Nature, 532(7598), 250-254.
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8

Immunohistochemical Analysis of Lung Tissue

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Disected lungs were subjected to 10% formalin fixation followed by paraffin embedment. Lung slices were stained with hematoxyline and eosin (H&E) or hybrisdized with antibodyes. In breif, lung slices were first rehydrated and treated with Hydrogen Peroxide Block (ThermoFisher) and then Protein Block (ThermoFisher). After blocking, the slices were incubated with primary antibody against GDF-15 (Sigma-Aldrich) overnight. The slices were then subjected to secondary Antibody Enhancer and Polymer-HRP (GBI LABS), followed by using diaminobenzene (ThermoFisher) as the chromogen before being mounted.
Chen Y.C., Wu C.T., Chen J.H., Tsai C.F., Wu C.Y., Chang P.C, & Yeh W.L. (2022). Diltiazem inhibits breast cancer metastasis via mediating growth differentiation factor 15 and epithelial-mesenchymal transition. Oncogenesis, 11(1), 48.
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9

Immunohistochemical Analysis of CLIC4, p53, TGF-β, TNF-α, and α-SMA

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For immunohistochemical analysis, 3-µm thick tissue sections were mounted on organosilane-coated slides (3-aminopropyltriethoxysilane; Sigma Chemical Co., St. Louis, MO, USA). Deparaffinization, rehydration and antigen retrieval were performed using Trilogy (Cell Marque, CA, USA) diluted in distilled water (1:100) and heated in a Pascal pressure cooker. Endogenous peroxidase and nonspecific antibody reaction were blocked with 3% hydrogen peroxide and Protein Block (Thermo Scientific, Runcorn, UK), respectively. The tissues were incubated with primary antibodies anti-CLIC4 (EPR14253, Abcam, 1:4000, 60'), anti-p53 (DO7, DAKO, 1:400, 60'), anti-TGF-β (3C11, Santa Cruz, 1:500, 60'), anti-TNF-α (52B83, Santa Cruz, 1:400, overnight) and anti-α-SMA (1A4, DAKO, 1:800, 60'). Antibodies were detected using the HiDef DetectionTM HRP Polymer system (Cell Marque) and reaction was developed with diaminobenzidine as chromogen (DAB, Sigma Chemical, St Louis, MO, USA). The sections were counterstained with Mayer's hematoxylin and mounted in Permount® (Fisher Scientific, Fair Lawn, NJ, USA). Replacement of the primary antibodies with bovine serum albumin was used as negative control and human melanoma tissue served as positive control.
Lima F.J., Lopes M.L.D.S., Barros C.C.D.S., Nonaka C.F.W, & Silveira É.J.D.D. (2020). Modification in CLIC4 Expression is Associated with P53, TGF-β, TNF-α and Myofibroblasts in Lip Carcinogenesis. Brazilian dental journal, 31(3).
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10

Immunohistochemical Staining of Mouse Tumors

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Mouse tumors were paraffin embedded and sectioned. Paraffin embedded sections were rehydrated through a xylene and alcohol series, rinsed in H2O and washed in PBS. The slides were then fixed with 10% formalin for 10 min and washed in PBS. Antigen retrieval was performed using target retrieval buffer (Vector Labs, Burlingame, CA) and steamed for 20 min. Samples were then blocked in a peroxidase blocking buffer (Thermo Scientific) for 15 min, followed by Protein block (Thermo Scientific) for 5 min, and incubated in appropriate primary antibody diluted in antibody diluent (S0809, Dako) at 4°C overnight in a humidified chamber. Following washing in PBS, samples were incubated in biotinylated anti-rabbit or polyvalent secondary antibody (Thermo Scientific) followed by streptavidin-HRP solution at room temperature for 20 min. Samples were then washed in PBS and incubated in 3-Amino-9-Ethyl-l-Carboazole (AEC) chromogen for up to 10 min. Finally, samples were washed in H2O, incubated in Mayers hematoxylin for 1 min, rinsed in cold H2O, and mounted in Aquamount.
Webster M.R., Fane M.E., Alicea G.M., Basu S., Kossenkov A.V., Marino G.E., Douglass S.M., Kaur A., Ecker B.L., Gnanapradeepan K., Ndoye A., Kugel C., Valiga A., Palmer J., Liu Q., Xu X., Morris J., Yin X., Wu H., Xu W., Zheng C., Karakousis G.C., Amaravadi R.K., Mitchell T.C., Almeida F.V., Xiao M., Rebecca V.W., Wang Y.J., Schuchter L.M., Herlyn M., Murphy M.E, & Weeraratna A.T. (2019). Paradoxical role for wild type p53 in driving therapy resistance in melanoma. Molecular cell, 77(3), 633-644.e5.
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