The immunofluorescence (IF) assay was performed on RAW264.7 cells as described previously [12 (link), 48 (link)]. Briefly, the primary antibodies against PCNA (1:50 dilution, Proteintech, USA), Rab7 (1:50 dilution, Proteintech, USA), LAMP1 (1:50 dilution, Proteintech, USA), LC 3 (1:100 dilution, Cell Signaling Technology, USA) and secondary antibodies conjugated with Alexa Fluor 488 and Alexa Fluor 594 dyes (Cat. no. ZF-0513, ZF-0511, and ZF-0512; ZSGB-BIO, China) were used for the test. To observe the superstructures of F-actin, phalloidine conjugated with Alexa Fluor 594 (Sigma, USA) was used to stain the cells for 1 hour at room temperature. The morphology of cell nuclei was shown by DAPI (Sigma, USA) staining for 10 min. The imaging experiments were digitized on laser scanning confocal microscopes (LSM700, Zeiss, Jena, Germany) as described previously [12 (link), 48 (link)]. In addition, the z-stacks plug-in was used to perform the consecutive scans of the cells in the direction of Z-axis.
Alexa fluor 594
Manufactured by ZSGB-BIO
Sourced in China
Alexa Fluor 594 is a fluorescent dye that can be used in various laboratory applications. It has an excitation maximum at 590 nm and an emission maximum at 617 nm. The dye is designed to provide bright, photostable fluorescence.
Automatically generated - may contain errors
Lab products found in correlation
Alexa fluor 488, by ZSGB-BIO (2 mentions)
4 6 diamidino 2 phenylindole (dapi), by Beyotime (2 mentions)
Lsm 700, by Zeiss (2 mentions)
Proliferating cell nuclear antigen (pcna), by Proteintech (1 mentions)
Alexa fluor 594, by Merck Group (1 mentions)
Lamp1, by Proteintech (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions)
Sp8 confocal microscope, by Leica (1 mentions)
Antifade mounting medium, by Beyotime (1 mentions)
3 protocols using alexa fluor 594
1
Immunofluorescence Assay of RAW264.7 Cells
2
Indirect Immunofluorescence Analysis of ACE2 and CD26
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For indirect immunofluorescence analysis, HEK293T cells were pre-seeded in a 15 mm culture dish and transfected with plasmids containing either eGFP-tagged hACE2 or hCD26. 24 h later, the cells were washed three times with PBS, fixed with 4% paraformaldehyde in PBS for 10 min, washed three times with PBS, and then blocked in PBS containing 1% bovine serum albumin for 1 h. The cells were then incubated with concentrated supernatant containing indicated proteins or purified MERS-RBD-mFc proteins (10 μg/mL). Cells were then washed three times with PBS and incubated with goat anti-mIgG conjugated with Alexa Fluor 594 (1:200, ZSGB-BIO) at room temperature for 1 h. Nuclei were stained with DAPI (5 μg/mL, Beyotime). The cells were then visualized on a Leica SP8 confocal microscope.
3
Immunofluorescence Staining of BV2 Cells
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BV2 cells cultured on glass coverslips were washed with PBS and xed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. 4 µm para n sections or BV2 cells were blocked with 5% goat serum albumin, and incubated with indicated primary antibodies overnight at 4°C. After incubation with secondary uorescent antibodies Alexa Fluor® 488 or Alexa Fluor® 594 antibodies (ZSGB-BIO, Beijing, China) for 1 h in the dark, the nucleuses were stained with DAPI (Beyotime). The coverslips were mounted onto glass slides using Antifade Mounting Medium (Beyotime), and the imaging was performed using NEXCOPE microscope (NE900, USA).
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