Alexa Fluor™ 594-labeled phalloidin (A12381) and Pierce™ BCA Protein Assay Kit (23227) were purchased from Thermo Fisher Scientific, China. 4′,6-Diamidino-2-phenylindole dihydrochloride (DAPI, D9542), PKH67 Green Fluorescent Cell Linker Kit for General Cell Membrane Labeling (MINI67-1KT), lipopolysaccharide (LPS, Escherichia coli 0111: B4, L4391), β‐glycerophosphate (G9422), L‐ascorbic acid 2‐phosphate (49752), and dexamethasone (D1756) were from Sigma, China. Antibodies used in this study were rabbit polyclonal antibody against alkaline phosphatase (ALP) (ab126820; Abcam, China) and CD81 antibody (B-11) (sc-166029; Santa Cruz Biotechnology, China).
Ab126820
Manufactured by Abcam
Sourced in United Kingdom
Ab126820 is a recombinant monoclonal antibody that specifically recognizes the protein of interest. The antibody is suitable for use in various laboratory applications, such as Western Blot and ELISA.
Automatically generated - may contain errors
Lab products found in correlation
Dexamethasone d1756, by Merck Group (1 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (1 mentions)
A10667, by Thermo Fisher Scientific (1 mentions)
H3570, by Thermo Fisher Scientific (1 mentions)
A11036, by Thermo Fisher Scientific (1 mentions)
Ht5011, by Merck Group (1 mentions)
Ab22552, by Abcam (1 mentions)
Ab90395, by Abcam (1 mentions)
Ab32084, by Abcam (1 mentions)
Ab58655, by Abcam (1 mentions)
A9434, by Merck Group (1 mentions)
Dp72 color digital camera, by Olympus (1 mentions)
A3059, by Merck Group (1 mentions)
Dm750, by Leica (1 mentions)
Alexa fluor 488 donkey anti mouse igg, by Thermo Fisher Scientific (1 mentions)
3 protocols using ab126820
1
Fluorescence Labeling and Protein Quantification
2
Immunofluorescence staining of cultured cells
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Cultured cell samples were carefully rinsed with PBS (Gibco, 21600-10), fixed with formalin solution (Sigma-Aldrich, HT5011, Madrid, Spain) for 20 min at room temperature, and rinsed again twice with PBS. The aldehyde groups were blocked with ammonium chloride (Sigma-Aldrich, A9434) 50 mM in PBS for 20 min. The samples were permeabilized with saponin (Sigma-Aldrich, 47036) 0.1% m/v in bovine serum albumin (BSA) (Sigma-Aldrich, A3059) 1% m/v in PBS for 10 min.
The samples were stained with the corresponding primary antibodies against paxillin (Abcam, ab32084, Cambridge, United Kingdom), scleraxis (Abcam, ab58655), collagen I (Abcam, ab90395), osterix (Abcam, ab22552), and alkaline phosphatase (Abcam, ab126820) in BSA 1% m/v PBS for 2 h at room temperature, then with the corresponding secondary fluorophore-conjugated antibodies anti-rabbit Alexa 568 (LifeTech, A11036, Madrid, Spain) and anti-mouse Alexa 488 (LifeTech, A10667) in BSA 1% m/v in PBS for 2 h. CytoPainter 488 (Abcam, ab176753) and Hoechst 33342 (Invitrogen, H3570) were used for actin filaments and cell nuclei staining. The samples were mounted with coverslips in fluoromount mounting medium (Sigma-Aldrich, HT5011).
The samples were imaged at a Nikon E600 upright manual microscope with a 40X/0.75 NA objective and an Olympus DP72 color digital camera. At least three representative images were taken of each sample.
The samples were stained with the corresponding primary antibodies against paxillin (Abcam, ab32084, Cambridge, United Kingdom), scleraxis (Abcam, ab58655), collagen I (Abcam, ab90395), osterix (Abcam, ab22552), and alkaline phosphatase (Abcam, ab126820) in BSA 1% m/v PBS for 2 h at room temperature, then with the corresponding secondary fluorophore-conjugated antibodies anti-rabbit Alexa 568 (LifeTech, A11036, Madrid, Spain) and anti-mouse Alexa 488 (LifeTech, A10667) in BSA 1% m/v in PBS for 2 h. CytoPainter 488 (Abcam, ab176753) and Hoechst 33342 (Invitrogen, H3570) were used for actin filaments and cell nuclei staining. The samples were mounted with coverslips in fluoromount mounting medium (Sigma-Aldrich, HT5011).
The samples were imaged at a Nikon E600 upright manual microscope with a 40X/0.75 NA objective and an Olympus DP72 color digital camera. At least three representative images were taken of each sample.
3
Histological Tissue Processing and Staining
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The fixed constructs (after formalin fixation as mentioned above) were subjected to standard histological tissue processing for paraffin embedding (EC350-2, Microm, Thermo Scientific, Waltham, MA, USA). Sections of 5 μm thickness were obtained using a microtome (HM355S, Microm, Thermo scientific). The sections were stained with hematoxylin-eosin (H&E) (05-12011/L, 05-M10003, Bio-optica) following a standard protocol and using an automatic stainer equipment (HMS740, Microm, Thermo Scientific).
For collagen, the sections were stained with Masson Trichrome (with aniline blue) assay kit (04-010802, Bio-Optica Milano S.p.A.) following manufacture's protocol.
For IHC, mouse IgG monoclonal antibody for Alkaline Phosphatase (ab126820, Abcam, UK, at 1:100 dilution) was used. Detection was done by using Alexa Fluor® 488 donkey anti-mouse IgG (A21202, Invitrogen, CA, at 1: 500 dilution) and counterstained with DAPI.
All sections after staining were dehydrated and mounted with Entellan® (4111, Inopat) before being observed under a transmitted and reflected light microscope (Leica DM750, Germany) with an MRc5 camera.
For collagen, the sections were stained with Masson Trichrome (with aniline blue) assay kit (04-010802, Bio-Optica Milano S.p.A.) following manufacture's protocol.
For IHC, mouse IgG monoclonal antibody for Alkaline Phosphatase (ab126820, Abcam, UK, at 1:100 dilution) was used. Detection was done by using Alexa Fluor® 488 donkey anti-mouse IgG (A21202, Invitrogen, CA, at 1: 500 dilution) and counterstained with DAPI.
All sections after staining were dehydrated and mounted with Entellan® (4111, Inopat) before being observed under a transmitted and reflected light microscope (Leica DM750, Germany) with an MRc5 camera.
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