The BK channel was fluorescently tagged through an immunofluorescence protocol with rabbit anti α-subunit antibody (APC107; Alomone Labs, Jerusalem, Israel), followed by Alexa-594 conjugated secondary antibody (Jackson ImmunoResearch, West Grove, PA). Briefly, neurons were incubated for 10-min with cold cholera toxin subunit-B conjugated to Alexa-488 (Life Technologies, Grand Island, NY). Dishes were fixed with cold 4% PFA (Electron Microscopy Sciences, Hatfield, PA) 4% sucrose (Sigma, St. Louis, MO) for 15-min. Permeabilization was then performed with 0.05% saponin (Sigma, St. Louis, MO). Neuronal dishes were blocked for 30-min RT with 5% BSA, 5% NDS in 1×PBS Na Azide and 0.005% saponin, followed by overnight incubation with primary antibody in 1/10 blocking solution at 4°C. The secondary was incubated for 1-hr RT in 1/10 blocking solution.
Apc 107
Manufactured by Alomone
Sourced in Palestine, State of, Israel
APC-107 is a laboratory equipment product designed for research purposes. It functions as a device for the analysis and detection of specific biomolecules.
Automatically generated - may contain errors
Lab products found in correlation
Alexa 594 conjugated secondary antibody, by Jackson ImmunoResearch (1 mentions)
Alexa 488, by Thermo Fisher Scientific (1 mentions)
Saponin, by Merck Group (1 mentions)
Sucrose, by Merck Group (1 mentions)
Paraformaldehyde (pfa), by Electron Microscopy Sciences (1 mentions)
Alexa fluor 488 goat anti rabbit igg, by Cell Signaling Technology (1 mentions)
Lsm710 confocal microscope, by Adobe (1 mentions)
Prolong gold antifade reagent, by Thermo Fisher Scientific (1 mentions)
Ab7817, by Abcam (1 mentions)
3 protocols using apc 107
1
BK Channel Immunofluorescence Labeling
2
Immunofluorescence Staining of Podocyte KCa1.1
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Podocytes were fixed in 4% paraformaldehyde in PBS for 15 min at room temperature. After rinsing twice with PBS, cells were blocked with 2% bovine serum albumen (BSA) in PBS for 1 h at room temperature. Cells on coverglass were incubated with KCa1.1 (1184–1200) antibody (APC-107, Alomone Labs) at a 1:100 dilution in PBS with 2% BSA at 4°C overnight. Cells were rinsed three times with PBS containing 0.02% Tween 20. Secondary antibodies were diluted in PBS with 2% BSA and cells were incubated at room temperature for 1 h. Alexa Fluor 488 goat anti-rabbit IgG (1:500; Cell Signaling Technology, United States) served as the secondary antibody. After rinsing three times with PBS containing 0.02% Tween 20, cells were incubated with DAPI (2 μg/ml) for 1 min. Cells were rinsed three times with PBS containing 0.02% Tween 20 and cells on coverglass were mounted on microscope slides with Prolong Gold antifade reagent (Invitrogen, United States). Images were taken using a Carl Zeiss LSM710 confocal microscope and processed using Photoshop software (Adobe Systems, Inc., San Jose, CA, United States). All of the images were quantified with ImageJ software and normalized to the respective control.
3
Immunofluorescence Staining of Rat Aortic Tissue
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Paraffin sections of the thoracic aorta of model rats were used for immunofluorescence staining.
After the sections were deparaffinized and hydrated, the antigen was repaired by the high-pressure method, blocked with 10% goat serum, and then incubated with primary antibodies against BKα (Alomone Labs, APC-107, Israel), BKβ1 (Alomone Labs, APC-036, Israel), CD3 (Abcam, ab5690, UK), CD19 (Bioss, bs-0079R, China), CD68 (Affinity Biosciences., DF7518, USA) and α-SMA (Abcam, ab7817, UK) at 4°C overnight. VSMCs were fixed in 4% paraformaldehyde for 10 min at room temperature. After washing with PBS, the smooth muscle cells were also blocked with 10% goat serum. Finally, the cells were incubated in NEDD4L and α-SMA primary antibodies at 4°C overnight.
The next day, the paraffin sections and cells were washed with PBS and incubated with a fluoresceinconjugated secondary antibody for 1 h at 37°C. After thorough washing, the tablets were sealed with a sealing solution containing DAPI. The image was acquired with a confocal microscope.
After the sections were deparaffinized and hydrated, the antigen was repaired by the high-pressure method, blocked with 10% goat serum, and then incubated with primary antibodies against BKα (Alomone Labs, APC-107, Israel), BKβ1 (Alomone Labs, APC-036, Israel), CD3 (Abcam, ab5690, UK), CD19 (Bioss, bs-0079R, China), CD68 (Affinity Biosciences., DF7518, USA) and α-SMA (Abcam, ab7817, UK) at 4°C overnight. VSMCs were fixed in 4% paraformaldehyde for 10 min at room temperature. After washing with PBS, the smooth muscle cells were also blocked with 10% goat serum. Finally, the cells were incubated in NEDD4L and α-SMA primary antibodies at 4°C overnight.
The next day, the paraffin sections and cells were washed with PBS and incubated with a fluoresceinconjugated secondary antibody for 1 h at 37°C. After thorough washing, the tablets were sealed with a sealing solution containing DAPI. The image was acquired with a confocal microscope.
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