ALI cultures treated with EGF for 24 h were fixed in a 50:50 mixture of methanol-acetone, permeabilized with 0.3% triton X-100, and blocked with 5% skimmed milk. Specimens were incubated overnight at 4 °C with TMEM16A antibody (1:200, Abcam) and MUC5AC antibody (1:200, Abcam), and then further incubated with secondary antibody rhodamine-conjugated goat anti-mouse IgG (1:500, Invitrogen) and fluorescein isothiocyanate-conjugated goat anti-rabbit IgG (1:500, Invitrogen). All specimens were then counterstained with 4, 6-diamidino-2-phenylinodole nuclear stain and examined by microscopy using an Olympus IX 81 confocal microscope (Tokyo, Japan). The number of TMEM16A-positive cells, MUC5AC-positive cells, and cells coexpressing TMEM16A and MUC5AC in 10 random fields of vision per culture was counted and expressed at a percentage of total cells.
Fluorescein isothiocyanate conjugated goat anti rabbit igg
Manufactured by Thermo Fisher Scientific
Sourced in United States
Fluorescein isothiocyanate-conjugated goat anti-rabbit IgG is a secondary antibody that binds to rabbit primary antibodies. It is conjugated with the fluorescent dye fluorescein isothiocyanate, enabling detection and visualization of the bound primary antibody.
Automatically generated - may contain errors
Lab products found in correlation
Ix81 confocal microscope, by Olympus (1 mentions)
Tmem16a antibody, by Abcam (1 mentions)
Muc5ac antibody, by Abcam (1 mentions)
Rhodamine conjugated goat anti mouse igg, by Thermo Fisher Scientific (1 mentions)
Anti pparα, by Proteintech (1 mentions)
Anti cpt1a, by Proteintech (1 mentions)
Anti srebp 1c, by Proteintech (1 mentions)
Deltavision elite, by GE Healthcare (1 mentions)
Anti fasn, by Santa Cruz Biotechnology (1 mentions)
Hoechst 33342, by Merck Group (1 mentions)
Anti cd81, by Abcam (1 mentions)
Axio scope a1 microscope, by Zeiss (1 mentions)
4 6 diamidino 2 phenylindole dihydrochloride, by Vector Laboratories (1 mentions)
3 protocols using fluorescein isothiocyanate conjugated goat anti rabbit igg
1
Analysis of TMEM16A and MUC5AC Expression in ALI Cultures
2
Immunofluorescence Analysis of Hepatic Lipid Metabolism
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Liver tissue and cultured cells were fixed in 4% paraformaldehyde (>30 min, at 4 °C) and then ruptured with 0.1% Triton X-100 (dilution in PBS) and blocked with 5% bovine serum albumin. Liver tissue segments and cultured cells were incubated with anti-CAMKK1 (ABclonal; rabbit, polyclonal, 1:100), anti-p-AMPK (Bioworld; rabbit, polyclonal, 1:200), anti-PPARα (Proteintech; rabbit, polyclonal, 1:100), anti-CPT-1A (Proteintech; rabbit, polyclonal, 1:100), anti-SREBP-1C (Proteintech; rabbit, polyclonal, 1:100), anti-FASn (Santa Cruz; mouse, monoclonal, 1:200), anti-p-IRS1 (Invitrogen; rabbit, polyclonal, 1:100), and anti-CD81 (Abcam; mouse, monoclonal, 1:100). Then the slides were labelled with fluorescein isothiocyanate-conjugated goat anti-rabbit IgG (Invitrogen; 1:200) and Alexa Fluor 555-conjugated donkey anti-mouse IgG. The nuclei were stained with Hoechst33342 (Sigma; 1:200). The slides were observed under a confocal microscope (DeltaVision Elite, GE, Boston, MA, USA), and the images were analysed using ImageJ 7.0 software.
3
Immunofluorescence Staining of Nuclear Protein
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Cells were fixed with 4% paraformaldehyde for 10 min and permeabilized with 0.1%Triton X-100 in PBS for a further 8 min, followed by blocking with 5% BSA/PBS for 1 h. Afterward, the cells were incubated with rabbit anti-p65 (1:100 dilution) at 4°C overnight, then incubated with fluorescein isothiocyanate-conjugated goat anti-rabbit IgG (Invitrogen) in the dark for 1 h. Coverslips were then mounted using a mounting medium with 4',6-diamidino-2-phenylindole dihydrochloride (Vector Laboratories, Burlingame, CA, USA). Effects were observed and photographed using an Axio Scope A1 microscope (Zeiss, Göttingen, Germany).
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