Mature CsgA fibers were deposited onto poly-L-lysine treated microscope slides51 (link) and slides were subsequently blocked for nonspecific binding by incubation with 5% (w/v) bovine serum albumin (BSA) for 10 min. The slides were then incubated at room temperature for 5 min with 3 μL 25 µM CsgC labeled with Alexa Fluor®488. Next, 10 μL of a mixture of an anti-6xHis antibody (AbD Serotec; MCA1396; 1:50 dilution) and an Alexa Fluor® 594-labeled goat anti-mouse antibody (Invitrogen; A11005; 1:20 dilution) in PBS was added for the staining of CsgA fibers. After 10 min incubation, slides were washed by flushing 3 times with 5 mL deionized water before examining them using a TE2000-U Nikon microscope.
Alexa fluor 594 labeled goat anti mouse antibody
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Alexa Fluor® 594-labeled goat anti-mouse antibody is a fluorescently-labeled secondary antibody designed for use in various immunodetection techniques. The Alexa Fluor® 594 dye provides a bright and photostable fluorescent signal that can be detected using standard red fluorescent filters.
Automatically generated - may contain errors
Lab products found in correlation
710 confocal microscope, by Zeiss (3 mentions)
Mca1396, by Bio-Rad (2 mentions)
Sp2 aobs confocal laser scanning microscope, by Leica (1 mentions)
Mab5406, by Merck Group (1 mentions)
Alexa fluor 488 labeled goat anti rabbit, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 488 labeled goat anti rabbit antibody, by Thermo Fisher Scientific (1 mentions)
Primary mouse antibody, by Santa Cruz Biotechnology (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Vector Laboratories (1 mentions)
4 protocols using alexa fluor 594 labeled goat anti mouse antibody
1
Visualizing CsgA Fibers with Fluorescent Labeling
2
Dual-label Immunofluorescence of FosB/ΔFosB
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Sections were rinsed and blocked 1 h in PBS containing 5% normal goat serum and 0.1% Tx in case of calcium/calmodulin-dependent protein kinase II (CaMKII) staining, or 0.02% Tx in the case of glutamic acid decarboxylase 67 (GAD67) staining. Sections were then incubated O/N at 4°C with rabbit anti-FosB/ΔFosB primary antibody (H-75, sc-7203, Santa Cruz Biotechnology; 1:250) and either mouse anti-CaMKII primary antibody (MA1–048; Pierce Biotechnology; 1:100) or mouse anti-GAD67 primary antibody (MAB5406; Chemicon International; 1:200). After washing, sections were incubated for 3 h with secondary antibodies Alexa Fluor 488-labeled goat anti-rabbit (A-11001; Invitrogen, Carlsbad, CA, USA; 1:500) and Alexa Fluor 594-labeled goat anti-mouse antibody (A-11005; Invitrogen, Carlsbad, CA, USA; 1:500). Sections were mounted and coverslipped with Mowiol. All fluorescent images were acquired using a Leica SP2 AOBS laser scanning confocal microscope (Leica Microsystems, Milan, Italy). Images for colocalization were captured at 40X magnification and the number of double-labeled cells were counted using Fiji software (http://fiji.sc/Fiji ).
3
Confocal Imaging of 14-3-3 in Rat Cardiomyocytes
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Confocal microscopy was used to detect 14-3-3 expression in adult rat cardiomyocytes as described previously 21 (link). Briefly, adult rat cardiomyocytes were isolated and plated on laminin-coated 4-well chamber slides (Lab-Tek). Expression of the 14-3-3 protein was detected using a primary mouse antibody (Santa Cruz, 1:100), and α-sarcomeric actinin was detected using a rabbit antibody (Sigma-Aldrich, 1:100). The secondary antibodies used were Alexa Fluor 594-labeled goat anti-mouse antibody and Alexa Fluor 488-labeled goat anti-rabbit antibody (Invitrogen, 1:500), and the mounting medium contained DAPI (Vector Laboratories). A Carl Zeiss 710 confocal microscope with ZEN software was used for visualizing 14-3-3, α-actinin, and DAPI. Total laser intensity and photomultiplier gain were constant for all the groups and settings, and data were verified by two independent observers who were blinded to the experimental group. A minimum of three coverslips was used for each experimental group, and at least three cell images were acquired from each coverslip.
4
Visualizing CsgA Fibers with Fluorescent Labeling
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Mature CsgA fibers were deposited onto poly-L-lysine treated microscope slides51 (link) and slides were subsequently blocked for nonspecific binding by incubation with 5% (w/v) bovine serum albumin (BSA) for 10 min. The slides were then incubated at room temperature for 5 min with 3 μL 25 µM CsgC labeled with Alexa Fluor®488. Next, 10 μL of a mixture of an anti-6xHis antibody (AbD Serotec; MCA1396; 1:50 dilution) and an Alexa Fluor® 594-labeled goat anti-mouse antibody (Invitrogen; A11005; 1:20 dilution) in PBS was added for the staining of CsgA fibers. After 10 min incubation, slides were washed by flushing 3 times with 5 mL deionized water before examining them using a TE2000-U Nikon microscope.
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