Rabbit anti rfp

Manufactured by Takara Bio

Sourced in United States

Rabbit anti-RFP is a primary antibody that specifically binds to the red fluorescent protein (RFP). It is used as a research tool to detect and localize RFP-tagged proteins in various applications, such as fluorescence microscopy and Western blotting.

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Anti-RFP (2 citations)

9 protocols using rabbit anti rfp

Immunohistochemical Staining Antibody Panel

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All experiments were performed as previously described [4 (link)]. A minimum of 5 animals has been analyzed for each combination of vectors. We used the following antibodies: rabbit anti-GFP (1:2000, Molecular Probe), chicken anti-GFP (1:2000, Abcam), rabbit anti-RFP (1:1000, Clontech), rabbit anti-5HT (1:10000, Incstar), mouse anti-TH (1:2000, RBI), mouse anti-NF200 (1:10000, Sigma), rabbit anti-ßgal (1:10000, CAPPEL), rabbit anti-Pax2 (1:4000, Abcam) and guinea-pig anti-PKCγ (1:7000, Strategic Biosolutions).

Bráz J.M., Wang F, & Basbaum A.I. (2015). Presynaptic Inputs to Any CNS Projection Neuron Identified by Dual Recombinant Virus Infection. PLoS ONE, 10(10), e0140681.

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Antibody Staining and Immunohistochemistry

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Antibody staining and immunohistochemistry were performed as previously described (Wang et al., 2004 (link)). The following primary antibodies were used: rabbit anti-GFP (Invitrogen, Carlsbad, CA, 1:1,000), mouse anti-GFP (Invitrogen, Carlsbad, CA, 1:1,000), mouse anti-nc82 (Hybridoma Bank, Iowa City, IA, 1:500), rabbit anti-RFP (Clontech, Mountain View, CA, 1:500); rabbit anti-GABA (Sigma-Aldrich, St. Louis, MO, 1:1,000); rabbit anti-FruM (1:100). For GRASP experiments, we used a mouse monoclonal antibody that specifically recognizes reconstituted GFP (Sigma-Aldrich, St. Louis, MO, 1:200). Secondary antibodies were Alexa Fluor goat anti-mouse 488, goat anti-rabbit 488, goat anti-mouse 568, goat anti-rabbit 568 (Life Technologies, Carlsbad, CA, 1:100).

Kallman B.R., Kim H, & Scott K. (2015). Excitation and inhibition onto central courtship neurons biases Drosophila mate choice. eLife, 4, e11188.

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Dual Immunofluorescence Histochemistry Protocol

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Brains were sectioned on a freezing microtome at 30 μm. Dual immunofluorescence histochemistry for cFOS-IR, mCherry-IR, and GFP-IR was conducted as previously described (Garfield et al., 2012 (link)). Primary antibodies (1/1,000) were as follows: rabbit anti-cFOS (EMD Millipore), rabbit anti-RFP (Clontech Laboratories, Inc.), or chicken anti-GFP (EMD Millipore).

Garfield A.S., Shah B.P., Madara J.C., Burke L.K., Patterson C.M., Flak J., Neve R.L., Evans M.L., Lowell B.B., Myers MG J.r, & Heisler L.K. (2014). A Parabrachial-Hypothalamic Cholecystokinin Neurocircuit Controls Counterregulatory Responses to Hypoglycemia. Cell Metabolism, 20(6), 1030-1037.

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Antibody Detection in AGS Cells

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Antibodies used in this study included rabbit anti-phospho-p38 (Thr180/Thr182, #9211, Cell Signaling Technology, Danver, MA, USA), rabbit anti-p38 (#9212, Cell signaling), mouse anti-GAPDH (MAB374) (Merck Millipore, Burlington, MA, USA), mouse anti-GMP1(SUMO-1) (18-2306, Life Technologies), rabbit anti-SUMO-2/3 (BML-PW9465, Enzo Life Sciences, Farmingdale, NY, USA), rabbit anti-lamin A/C (#2032, Cell Signaling), rabbit anti-RFP (632496, Clontech, Mountain View, CA, USA), rabbit anti-GST (71-7500, Life Technologies), rabbit anti-His (SC-803, Santa Cruz Biotechnology, Dallas, TX, USA), horseradish peroxidase (HRP)-conjugated goat anti-mouse IgG (G21040, Life Technologies) and HRP-conjugated goat anti-rabbit IgG (G21234, Life Technologies). The rabbit anti-phospho-p38 and the rabbit anti-p38 used in this study can both react with the various isoforms of p38 (p38α, p38β, p38γ, and p38δ). However, the main isoform expressed in AGS cells is p38α, so this is referred to simply as p38 throughout the paper.

Wang P.Y., Hsu P.I., Wu D.C., Chen T.C., Jarman A.P., Powell L.M, & Chen A. (2018). SUMOs Mediate the Nuclear Transfer of p38 and p-p38 during Helicobacter Pylori Infection. International Journal of Molecular Sciences, 19(9), 2482.

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Immunostaining of Larval and Adult Drosophila Brains

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Larval and Adult brains were fixed in 4% formaldehyde for 20 min and stained as previously described^{51 (link)}. The following primary antibodies were used: rabbit anti-GFP 1:1000 (LifeTechnologies), mouse anti-Acj6 1:10 (DSHB), mouse anti-Elav 1:20 (9F8A9, DSHB), rat anti-Elav 1:20 (7E8A10, DSHB), mouse anti-Ey 1:20 (DSHB) mouse anti-Dac 1:40 (mAbdac2–3, DSHB), rat anti-CadN 1:20 (DN-Ex #8, DSHB), mouse anti-Chp 1:50 (24B10, DSHB), mouse anti-Fas2 1:10 (1D4, DSHB), rabbit anti-RFP 1:500 (clontech), mouse anti-Repo 1:20 (8D12, DSHB), rabbit anti-Mira 1:500, rabbit anti-pH3 1:200 (Millipore), mouse anti-Dlg 1:50 (4F3, DSHB), guinea pig anti-Dpn 1:500 (Brand lab). DNA was stained using DAPI or TOPRO-3 1:200 (Molecular Probes, Invitrogen).
Alexa Fluor conjugated secondary antibodies were diluted 1:200 (Molecular Probes, Invitrogen). Primary and secondary antibodies were incubated at 4 °C overnight. Brains were mounted on slides in Vectashield (Vector).

Contreras E.G., Palominos T., Glavic Á., Brand A.H., Sierralta J, & Oliva C. (2018). The transcription factor SoxD controls neuronal guidance in the Drosophila visual system. Scientific Reports, 8, 13332.

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EdU Detection and Immunostaining Protocol

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We used the click-it® chemical reaction protocol for EdU detection followed by immunostaining with primary and secondary antibodies and before DAPI staining. For confocal microscopy, embryos were fixed in 4% paraformaldehyde overnight and then 30% sucrose, and then embedded in OCT, sectioned and stained using standard protocols. Antibodies used were: goat and rabbit anti-β1-integrin (1:400; R&D or 1:1000; Abcam), mouse anti-Isl1 (1:200, Iowa Hybridoma Bank), rabbit anti-RFP (1:400, Clontech), rabbit anti-Numb (pre-absorbed, 1: 500, Abcam or from Dr. Zhong), and mouse anti-PH3 (1:500, Abcam). Alexa Fluor secondary antibodies (Invitrogen) were used for all secondary detection and confocal images acquired with a Zeiss LSM 510 Meta confocal microscope using Zen™ acquisition software. For Western blotting, cell lysate was resolved on SDS-PAGE and electroblotted on nitrocellulose membranes and incubated with primary antibodies in 5% nonfat milk overnight at 4 degrees Celsius. Secondary antibodies were incubated for 1 hour at room temperature. The blots were washed 3×10 mins in TTBS, and detection was by chemiluminescence (Amersham ECL, GE Healthcare Life Sciences).

Gibbs B., Shenje L., Andersen P., Miyamoto M, & Kwon C. (2018). β1-integrin Is a Cell-Autonomous Factor Mediating the Numb Pathway for Cardiac Progenitor Maintenance. Biochemical and biophysical research communications, 500(2), 256-260.

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Midgut Immunostaining for Developmental Markers

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A dissected female adult midgut at an appropriate adult stage was fixed with 4% paraformaldehyde at room temperature for 40 min. After washing with phosphate-buffered saline containing 0.1% TritonX-100, immunostaining was carried out with the following primary antibodies: rat anti-GFP (Nacalai Tesque, 1:200 dilution), rabbit anti-RFP (Clontech, 1:200 dilution), chicken anti-beta-galactosidase (abcam, 1:200 dilution), rabbit anti-phospho-histone H3 (pH3) (Upstate Biotech, 1:200 dilution), mouse anti-Prospero (DSHB, 1:100 dilution), mouse anti-AstA (DSHB,1:10) (Woodhead et al., 1989) , mouse anti-Dl (DSHB, 1:100 dilution), and rabbit anti-Dh31 (1:500 dilution, a generous gift from Dr. Jan Veenstra) (Veenstra et al., 2008) . The secondary antibodies used were Alexa Fluor647 or Alexa Fluor555-conjugated anti-mouse IgG (Jackson Immuno Research, 1:200 dilution), Alexa488-conjugated anti-rat IgG (Jackson Immuno Research, 1:200 dilution), Alexa Fluor555 or Alexa Fluor647-conjugated anti-rabbit IgG (Jackson Immuno Research, 1:200), and DyLight649-conjugated anti-chicken IgY (Jackson Immuno Research, 1:200 dilution). Nuclei were stained with 4'6-diamidino-2-phenylindole (DAPI, SIGMA).

Takeda K., Okumura T., Terahata M., Yamaguchi M., Taniguchi K, & Adachi-Yamada T. (2018). Drosophila Peptide Hormones Allatostatin A and Diuretic Hormone 31 Exhibiting Complementary Gradient Distribution in Posterior Midgut Antagonistically Regulate Midgut Senescence and Adult Lifespan. Zoological science, 35(1).

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Quantification of Neuron Connectivity using GRASP

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Antibody staining was performed as previously described (Wang et al., 2004 (link)). The following primary antibodies were used: rabbit anti-GFP (Invitrogen, Carlsbad, CA 1:1000), mouse anti-GFP (Invitrogen 1:1000), mouse anti-GFP (Sigma-Aldrich, St. Louis, MO 1:200 - for GRASP staining only), chicken anti-GFP (1:500), mouse anti-nc82 (Developmental Studies Hybridoma Bank, Iowa City, IA 1:500), rabbit anti-RFP (Clonetech, Mountain View, CA 1:500). The following secondary antibodies were used (all Invitrogen at 1:100): 488 anti-rabbit, 488 anti-mouse, 488 anti-chicken, 568 anti-rabbit, 568 anti-mouse, 647 anti-mouse. All images were acquired on a Zeiss confocal microscope. Brightness and contrast were adjusted using FIJI. For quantification of GRASP, we used the following steps for each image: (1) We made a mask (M) of the TPN dendritic field using Huang threshold in FIJI; (2) Using the distribution of GFP outside the mask (M_out), we choose the 99th percentile as the threshold (T); (3) We measured the percent of pixels inside the mask (M) that exceed the threshold (T).

Kim H., Kirkhart C, & Scott K. (2017). Long-range projection neurons in the taste circuit of Drosophila. eLife, 6, e23386.

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Immunohistochemical Staining Protocol

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Immunohistochemical staining was performed as previously described [54 (link)]. The primary antibodies used were rabbit anti-GFP (Invitrogen, Carlsbad, CA 1:1000), mouse anti-GFP (Invitrogen 1:1000), mouse anti-nc82 (Developmental Studies Hybridoma Bank, Iowa City, IA 1:500), rabbit anti-RFP (Clonetech, Mountain View, CA 1:500); rabbit α-HA Tag (Cell Signaling Technologies 1:300), DL550 mouse α-V5 (Bio-Rad 1:500), chicken anti-GFP (Life Technologies, 1:500). The secondary antibodies were (all Invitrogen at 1:100): 488 anti-rabbit, 488 anti-mouse, 488 anti-chicken, 568 anti-rabbit, 568 anti-mouse. All images were acquired on a confocal microscope. Afterward, brightness and contrast were adjusted with FIJI (ImageJ) software.

Youn H., Kirkhart C., Chia J, & Scott K. (2018). A subset of octopaminergic neurons that promotes feeding initiation in Drosophila melanogaster. PLoS ONE, 13(6), e0198362.

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