biotinylated anti-mouse igg1 second-step reagent  (SouthernBiotech)

Journal: Bio-protocol

Article Title: An Integrated Workflow for Three-Dimensional Visualization of Human Skeletal Muscle Stem Cell Nuclei

doi: 10.21769/BioProtoc.5281

Figure Lengend Snippet: Immunohistochemistry antibodies and reagent information

Article Snippet: Biotinylated-SP goat anti-mouse IgG1 secondary antibody , Jackson ImmunoResearch , 115-065-205 , Mouse IgG1 , Goat/IgG1 , (1:1,000)/2.5% NHS , -20 °C.

Techniques: Immunohistochemistry

Journal: Bio-protocol

Article Title: An Integrated Workflow for Three-Dimensional Visualization of Human Skeletal Muscle Stem Cell Nuclei

doi: 10.21769/BioProtoc.5281

Figure Lengend Snippet: The C2C12 (ATCC CRL-1772) mouse myoblast cell line incubated in sp2/0-ag14 conditioned serum media was used as a negative control for Pax7 identification. The sp2/0-ag14 serum replaced the traditional horse serum, which induces differentiation and expression of Pax7. This figure plate shows a very weak nonspecific background signal but not Pax7 positive labeling. C2C12 cells were thawed at passage 13 (P13) and cultured for 5 days in Dulbecco’s modified Eagle’s medium + 1% penicillin-streptomycin + 10% heat-inactivated FBS at 37 °C in an incubator with 5% CO 2 . Cells were then passaged and incubated in the same culture media but with sp2/0-ag14 conditioned sera for 3 days. The immunohistochemical protocol was run at cell passage 14 (P14) in an 8-well Ibidi μ-slide with a 200 μL working volume. Briefly, cells were fixed in 2% formaldehyde for 10 min, blocked in 3% hydrogen peroxide for 7 min to quench endogenous peroxidase activity, blocked in 3% normal goat serum diluted in 1× HBSS+0.1% sucrose+1.0% saponin for 20 min, and incubated overnight in a mouse monoclonal anti-Pax7 IgG1 primary antibody (dilution 1:100). The next day, cells were incubated in goat anti-mouse IgG1 biotin-SP-conjugated secondary antibody (dilution 1:1,000) for 60 min, incubated in streptavidin horseradish peroxidase conjugate for 60 min, amplified with Alexa Fluor 594 tyramide conjugate (dilution 1:200) for 20 min, and counterstained with 10 mM DAPI for 30 min. VectaShield antifade mounting media was added, and plates were stored at 4°C until imaging. Images were acquired using an Olympus IX-81 epifluorescent inverted microscope with the Olympus UPlanSApo 40×/0.95 NA long-working-distance objective and SlideBook 6.0 software. Scale bar = 20 μm.

Article Snippet: Biotinylated-SP goat anti-mouse IgG1 secondary antibody , Jackson ImmunoResearch , 115-065-205 , Mouse IgG1 , Goat/IgG1 , (1:1,000)/2.5% NHS , -20 °C.

Techniques: Incubation, Negative Control, Expressing, Labeling, Cell Culture, Modification, Immunohistochemical staining, Activity Assay, Amplification, Imaging, Inverted Microscopy, Software

Journal: Bio-protocol

Article Title: An Integrated Workflow for Three-Dimensional Visualization of Human Skeletal Muscle Stem Cell Nuclei

doi: 10.21769/BioProtoc.5281

Figure Lengend Snippet: The C2C12 (ATCC CRL-1772) mouse myoblast cell line incubated in differentiation media containing horse serum was used as a positive control for Pax7 identification. Horse serum induces differentiation and expression of Pax7 when applied for more than two days. This figure plate shows a strong overlapping signal with DAPI (merged image), indicating similar signal intensities in both the muscle tissue samples and C2C12 cells. C2C12 cells were thawed at passage 12 (P12) and cultured for two weeks in Dulbecco’s modified Eagle’s medium + 1% penicillin-streptomycin + 10% heat-inactivated FBS at 37 °C in an incubator with 5% CO 2 . Cells were then passaged and incubated in differentiation culture media replacing FBS with horse serum for 3 days. The immunohistochemical protocol was run at cell passage 13 (P13) in an 8-well Ibidi μ-slide with a 200 μL working volume. Briefly, cells were fixed in 2% formaldehyde for 10 min, blocked in 3% hydrogen peroxide for 7 min to quench endogenous peroxidase activity, blocked in 3% normal goat serum diluted in 1× HBSS + 0.1% sucrose + 1.0% saponin for 20 min, and incubated overnight in a mouse monoclonal anti-Pax7 IgG1 primary antibody (dilution 1:100). The next day, cells were incubated in goat anti-mouse IgG1 biotin-SP-conjugated secondary antibody (dilution 1:1,000) for 60 min, incubated in streptavidin horseradish peroxidase conjugate for 60 min, amplified with Alexa Fluor 594 tyramide conjugate (dilution 1:200) for 20 min, and counterstained with 10 mM DAPI for 30 min. VectaShield antifade mounting media was added, and plates were stored at 4°C until imaging. Images were acquired using an Olympus IX-81 epifluorescent inverted microscope with the Olympus UPlanSApo 40×/0.95 NA long-working-distance objective and SlideBook 6.0 software. Scale bar = 20 μm.

Article Snippet: Biotinylated-SP goat anti-mouse IgG1 secondary antibody , Jackson ImmunoResearch , 115-065-205 , Mouse IgG1 , Goat/IgG1 , (1:1,000)/2.5% NHS , -20 °C.

Techniques: Incubation, Positive Control, Expressing, Cell Culture, Modification, Immunohistochemical staining, Activity Assay, Amplification, Imaging, Inverted Microscopy, Software

Journal: eLife

Article Title: Immunogenicity and safety of a live-attenuated SARS-CoV-2 vaccine candidate based on multiple attenuation mechanisms

doi: 10.7554/elife.97532

Figure Lengend Snippet: Figure 2. Immunogenicity of the vaccine candidate in hamsters. (A) Hamsters were inoculated with 1 × 103 or 1 × 104 plaque-forming unit (PFU) of BK2102 intranasally, and the serum was collected 4 weeks after inoculation. Spike-specific IgG in the sera of BK2102-inoculated hamsters and mock- treated hamsters was detected by ELISA. Symbols depict data of individual hamsters (n=10), and bars correspond to the median value. The limit of dilution is indicated in the x-axis. (B) Neutralizing antibodies in the sera were induced in BK2102-inoculated hamsters. Neutralizing antibodies in the

Article Snippet: Biotinylated anti- Syrian hamster IgG1 Ab (Southern Biotech, Cat# 1940- 08, 1/100 dilution), IgG2/3 subclass Ab (Southern Biotech, Cat# 1935- 08, 1/200,000 dilution), and IgA Ab (Brookwood Biomedical, Cat# 3003a, 1/100 dilution) were used and detected with HRPlabeled streptavidin (Abcam, Cat# ab7403, 1/10,000 dilution).

Techniques: Immunopeptidomics, Enzyme-linked Immunosorbent Assay