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Anti Brp (nc82) is a mouse monoclonal antibody that recognizes the Bruchpilot (Brp) protein, a key component of the active zone in presynaptic terminals. It is a useful tool for the identification and localization of Brp in various biological samples.

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Lab products found in correlation

Dapi fluoromount g, by Southern Biotech (2 mentions) Goat anti mouse alexa 488, by Thermo Fisher Scientific (2 mentions) Slowfade glass antifade mountant, by Thermo Fisher Scientific (2 mentions) Anti futsch 22c10, by Developmental Studies Hybridoma Bank (2 mentions) Goat anti mouse tritc, by Jackson ImmunoResearch (2 mentions) Anti rabbit fitc, by Jackson ImmunoResearch (2 mentions) Alexa 647 conjugated goat anti hrp, by Jackson ImmunoResearch (2 mentions) Mouse anti dlg 4f3, by Developmental Studies Hybridoma Bank (2 mentions) A1r resonant scanning confocal microscope, by Nikon (1 mentions) Nc 82, by Thermo Fisher Scientific (1 mentions)

4 protocols using anti brp nc82

1

Drosophila Larval Neuromuscular Junction Staining

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Third instar larvae were dissected in ice-cold 1xPBS and fillets were fixed in 4%PFA in PBS at room temperature for 30min. For Futsch staining, fillets were fixed in ice-cold methanol for 10min at −20°C. Samples were washed in 1xPBS for 10 min followed by two washes in 1xPBST (PBS + 0.1% Triton) for 10min each. Fillets were transferred to 0.5μl tubes for primary antibody incubation overnight at 4°C with agitation. After three 15 min washes in 1xPBST, samples were incubated in secondary antibodies for 1 h at room temperature. Samples were then washed three times for 10 min each and mounted with Fluoromount-G DAPI (SouthernBiotech) or SlowFade Glass Antifade Mountant (Invitrogen). Mouse anti-Dlg (4F3) 1:1000, anti-Futsch (22C10) 1:100, and anti Brp (nc82) 1:100 were obtained from Developmental Studies Hybridoma Bank (DSHB, University of Iowa). Rabbit anti- GluRIIC (1:5000) was a kind gift from Aaron DiAntonio. Alexa 647-conjugated goat anti-HRP was used at 1:200 (Jackson ImmunoResearch Laboratories, Inc., West Grove, PA). Goat anti-mouse Alexa 488 (Invitrogen) was used at 1:400 while goat anti-mouse TRITC and anti-rabbit FITC (Jackson ImmunoResearch Laboratories, Inc., West Grove, PA) were used at 1:400.
Belalcazar H.M., Hendricks E.L., Zamurrad S., Liebl F.L, & Secombe J. (2021). The histone demethylase KDM5 is required for synaptic structure and function at the Drosophila neuromuscular junction. Cell reports, 34(7), 108753.
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2

Drosophila NMJ Immunostaining and Analysis

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Drosophila larvae were reared and immunostained as described 16 (link). Wild type (w1118) or GR.100 larvae (w;OK6-Gal4/UAS-100xGR) were reared on individual apple plates at 25°C and fed yeast mixed with either solvent (DMSO) or apilimod (30 µM) for 5 days throughout larval development. Briefly, third-instar larvae were dissected in saline and immunostained with anti-BRP (nc82; Developmental Studies Hybridoma Bank; 1:100) and anti-HRP (Cyanine 3 (Cy3) conjugated; Jackson ImmunoResearch; 1:400). Alexa Fluor 488-conjugated mouse secondary antibody was used at 1:400 (Jackson ImmunoResearch). NMJs were imaged using a Nikon A1R Resonant Scanning Confocal microscope with a 100x APO 1.4 NA oil immersion objective. BRP puncta were quantified using Nikon Elements Software and data was analyzed using Graphpad Prism.
Hung S.T., Linares G.R., Chang W.H., Eoh Y., Krishnan G., Mendonca S., Hong S., Shi Y., Santana M., Kueth C., Macklin-Isquierdo S., Perry S., Duhaime S., Maios C., Chang J., Perez J., Couto A., Lai J., Li Y., Alworth S.V., Hendricks E., Wang Y., Zlokovic B.V., Dickman D.K., Parker J.A., Zarnescu D.C., Gao F.B, & Ichida J.K. (2023). PIKFYVE Inhibition Mitigates Disease in Models of Diverse Forms of ALS. Cell, 186(4), 786-802.e28.
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3

Immunostaining of Drosophila Neuromuscular Junctions

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Wandering third instar larvae were dissected in Ca2+-free HL3 solution and fixed in 4% paraformaldehyde for 10 min, washed in PBT (PBS plus 0.1% Triton X-100) and blocked in 5% normal goat serum (NGS) and 5% BSA in PBT for 15 min. Samples were incubated overnight with anti-BRP (NC82, 1:200) from the Developmental Studies Hybridoma Bank (DSHB Cat# NC82, RRID:AB_2314866), washed for 1 hr in PBS and then incubated for 2–3 hr with Alexa Fluor 607-conjugated anti-mouse IgG at 1:1000 (Invitrogen, #A21237, RRID:AB_1500743).
Akbergenova Y., Cunningham K.L., Zhang Y.V., Weiss S, & Littleton J.T. (2018). Characterization of developmental and molecular factors underlying release heterogeneity at Drosophila synapses. eLife, 7, e38268.
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4

Drosophila Larval Neuromuscular Junction Staining

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Third instar larvae were dissected in ice-cold 1xPBS and fillets were fixed in 4%PFA in PBS at room temperature for 30min. For Futsch staining, fillets were fixed in ice-cold methanol for 10min at −20°C. Samples were washed in 1xPBS for 10 min followed by two washes in 1xPBST (PBS + 0.1% Triton) for 10min each. Fillets were transferred to 0.5μl tubes for primary antibody incubation overnight at 4°C with agitation. After three 15 min washes in 1xPBST, samples were incubated in secondary antibodies for 1 h at room temperature. Samples were then washed three times for 10 min each and mounted with Fluoromount-G DAPI (SouthernBiotech) or SlowFade Glass Antifade Mountant (Invitrogen). Mouse anti-Dlg (4F3) 1:1000, anti-Futsch (22C10) 1:100, and anti Brp (nc82) 1:100 were obtained from Developmental Studies Hybridoma Bank (DSHB, University of Iowa). Rabbit anti- GluRIIC (1:5000) was a kind gift from Aaron DiAntonio. Alexa 647-conjugated goat anti-HRP was used at 1:200 (Jackson ImmunoResearch Laboratories, Inc., West Grove, PA). Goat anti-mouse Alexa 488 (Invitrogen) was used at 1:400 while goat anti-mouse TRITC and anti-rabbit FITC (Jackson ImmunoResearch Laboratories, Inc., West Grove, PA) were used at 1:400.
Belalcazar H.M., Hendricks E.L., Zamurrad S., Liebl F.L, & Secombe J. (2021). The histone demethylase KDM5 is required for synaptic structure and function at the Drosophila neuromuscular junction. Cell reports, 34(7), 108753.
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