High ph buffer

Manufactured by Vector Laboratories

High pH buffer is a laboratory solution designed to maintain a stable pH environment within the alkaline range. It is commonly used for various applications that require a consistent alkaline pH, such as protein purification, enzyme assays, and biological sample preparation.

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Lab products found in correlation

Af3626, by R&D Systems (1 mentions) Af3625, by R&D Systems (1 mentions) In situ cell death detection kit, by Roche (1 mentions) Alexa fluor 635 phalloidin, by Thermo Fisher Scientific (1 mentions) Vectashield with dapi, by Vector Laboratories (1 mentions) Anti goat alexafluor594, by Thermo Fisher Scientific (1 mentions) Alexa fluor 488 goat anti rabbit, by Jackson ImmunoResearch (1 mentions) Decloaking chamber, by Biocare Medical (1 mentions)

3 protocols using high ph buffer

Immunohistochemical Analysis of IL-33 and Proliferation

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Briefly, sections were deparaffinized, rehydrated, and antigen was unmasked in a high pH buffer (Vector Labs, Burlingame, CA). Sections were stained with goat anti-human or anti-mouse IL-33 antibodies (AF3625 and AF3626, R&D Systems, Minneapolis, MN). An isotype goat polyclonal IgG negative control was used to test for the specificity of the involved antibody at matched concentration and incubation conditions (Jackson Immunoresearch, West Grove, PA). Sections were counterstained with Mayer’s hematoxylin.
Sections were also stained with anti-Ki67 (790-4286, Ventana, Tuscon, AZ) or phospho-histone H3 (9701S, Cell signaling, Danvers, MA) to quantify proliferation. Apoptotic epithelial cells were identified based on positive TUNEL staining (In Situ Cell Death Detection Kit, Roche, Indianapolis, IN).

Waddell A., Vallance J.E., Moore P.D., Hummel A.T., Wu D., Shanmukhappa S.K., Fei L., Washington M.K., Minar P., Coburn L.A., Nakae S., Wilson K.T., Denson L.A., Hogan S.P, & Rosen M.J. (2015). IL-33 signaling protects from murine oxazolone colitis by supporting intestinal epithelial function. Inflammatory bowel diseases, 21(12), 2737-2746.

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Immunofluorescent Analysis of IL-33 and F4/80 in Colon Sections

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For immunofluorescent staining of IL-33 with F4/80 on colon sections, sections were deparaffinized, rehydrated, and antigen was unmasked in a high pH buffer (Vector Labs, Burlingame, CA). Sections were stained with goat anti-mouse IL-33 (AF3626, R&D Systems, Minneapolis, MN) followed by anti-goat-Alexa Fluor-594 (Life Technologies, Carlsbad, CA) with anti-F4/80-FITC (11-4801-11, eBioscience, San Diego, CA) and sections were counterstained with Vectashield with DAPI (Vector Laboratories, Burlingame, CA).
T84 cells grown on coverslips were subsequently fixed with 4% paraformaldehyde for 30 minutes. Cells were blocked with goat serum and then stained with rabbit anti-ST2 (ab25877, AbCam, Cambridge, MA) followed by goat anti-rabbit-Alexa Fluor488 (Jackson Immunoresearch, West Grove, PA), and Alexa Fluor635-Phalloidin (Life Technologies, Carlsbad, CA) and counterstained with Vectashield with DAPI (Vector Laboratories, Burlingame, CA).

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Immunohistochemical Analysis of CK17 and CK7 Expression

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Briefly, sections were cut at 4 μm and heat immobilized at 60°C for 60 minutes. After deparaffinization with xylene and rehydration through graded alcohols, antigen retrieval was performed in a High PH buffer (Vector Laboratories) at 120°C for 10 minutes in a Decloaking Chamber (Biocare Medical, Walnut Creek, CA). Sections were rinsed in phosphate-buffered saline (PBS) and blocked for 1 hour at room temperature using 5% nonfat milk. Primary antibodies were added and incubated overnight at 4°C using the following: monoclonal CK17 (E3 clone) prediluted (Abcam) and rabbit polyclonal CK7, diluted 1:1,000 (Abcam). Sections were rinsed in PBS and incubated with Alexa secondary 488 goat anti-mouse and 594 goat anti-rabbit, both diluted to 1:250, incubated for 2 hours at room temperature in a light-proof chamber. The slides were rinsed in PBS and cover slipped using Vectashield mounting media with 4',6-diamidino-2-phenylindole (Vector Laboratories). Negative controls were performed on all cases.

Mockler D., Escobar-Hoyos L.F., Akalin A., Romeiser J., Shroyer A.L, & Shroyer K.R. (2017). Keratin 17 Is a Prognostic Biomarker in Endocervical Glandular Neoplasia. American journal of clinical pathology, 148(3).

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