Rabbit anti pgp9

Manufactured by Abcam

Sourced in Japan, United States

Rabbit anti-PGP9.5 is a primary antibody used to detect the presence and distribution of the protein PGP9.5 in biological samples. PGP9.5 is a neuronal marker commonly used in immunohistochemistry and Western blot applications.

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Anti-PGP9.5 (6 citations) Rabbit anti- (5 citations)

5 protocols using rabbit anti pgp9

Immunohistochemical Staining of Tissue Sections

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Tissue sections were rinsed in PBS before adding blocking solution [PBS + 10% (v/v) goat serum (Sigma)/1% bovine serum albumin (Fisher Scientific)/0.3% Triton X-100 (Fisher)] for 1 h. Primary antibodies were then diluted in blocking solution and incubated overnight at 4 °C in a humidified slide staining chamber. Lyve1⁺ and MHCII⁺ were detected using rat anti-Lyve1 (Clone: ALY7;eBioscience; 1: 500) and rat anti-MHCII (Clone: M5/114.15.2; Novus Biologicals; 1: 500) primary antibodies, paired with rabbit anti-CD68 (polyclonal; Abcam; 1: 1000)/rabbit anti-Iba1 (polyclonal; Wako; 1: 1000), rabbit anti-CD68 (polyclonal; Abcam; 1: 500), rabbit anti-PGP9.5 (Clone: EPR4118; Abcam; 1: 200), rabbit anti-CD206/MRC1 (Clone E6T5J; Cell Signaling; 1: 500), or rabbit anti-CD31 (Clone: SP38; Invitrogen; 1: 50). T cells were detected with rat anti-CD3 (Clone: CD3-12; Abcam; 1: 100). Slides were then rinsed 10 × in PBS. Secondary antibodies (goat anti-rabbit Alexa Fluor-594 with goat anti-rat Alexa Fluor-488, or goat anti-rabbit Alexa Fluor-488 with goat anti-rat Alexa Fluor-594; 1: 500; Invitrogen) in PBS containing DAPI (500 ng/ml; Sigma) were placed on the slides and incubated for 3 h at RT. Sections were again rinsed 10 × in PBS and coverslipped with FluorSave mounting medium (Millipore).

Nichols J.M., Pham H.V., Lee E.F., Mahalingam R, & Shepherd A.J. (2024). Single-cell analysis of age-related changes in leukocytes of diabetic mouse hindpaws. Cellular and Molecular Life Sciences: CMLS, 81(1), 146.

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Immunolabeling of Neural Markers

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The following antibodies were used as primary antibodies for immunocytostaining and for immunohistochemistry: rabbit anti‐s100b (dilution = 1:1,000) (Dako, Ely, UK), rabbit anti‐p75 (1:500) (Cell Signaling Technology, Danvers, MA, USA), rabbit anti‐GAP43 (1:200) (Cell Signaling Technology), rabbit anti‐glial fibrillary acidic protein (GFAP) (1:200) (Abcam, Cambridge, UK), rabbit anti‐NG2 (1:1,000) (Millipore, Cambridge, UK), mouse anti‐myelin basic protein (MBP) (1:100) (Chemicon, Chandlers Ford, UK), goat anti‐protein zero (P0) (1:500) (Abcam), rabbit anti‐Tuj‐1 (1:100) (Chemicon), mouse anti‐neurofilament (NF) (1:1,000) (Sigma), rabbit anti‐PGP9.5 (1:500) (Abcam), mouse anti‐fibronectin (Millipore), mouse anti‐Nanog (1:200) (ReproCELL, Yokohama, Japan), mouse anti‐MAP‐2 (1:200) (Abcam), and rabbit anti‐NeuN (1:500) (Abcam) antibodies. As secondary antibodies, Alexa Fluor 546‐conjugated anti‐rabbit and anti‐mouse antibodies and Alexa Fluor 670‐conjugated anti‐rabbit and anti‐mouse antibodies (1:1,000) (Life technologies, Carlsbad, CA, USA) were used.

Sowa Y., Kishida T., Tomita K., Yamamoto K., Numajiri T, & Mazda O. (2017). Direct Conversion of Human Fibroblasts into Schwann Cells that Facilitate Regeneration of Injured Peripheral Nerve In Vivo. Stem Cells Translational Medicine, 6(4), 1207-1216.

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Immunohistochemical Analysis of Neural Markers

Cited 1 time

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Immunohistochemistry was performed as described^{19 (link)}. The following primary antibodies were used: Human anti-Anna-1 (Hu) positive serum (1:16,000, generous gift from Dr. Vanda Lennon; Mayo Clinic, Rochester MN;), goat anti-Sox2 (1:50; R&D Systems, Minneapolis, MN, Cat # AF2018), mouse anti-Tuj1 (1:100; Covance, Dedham, MA Cat # MMS-435P), goat anti-GFAP (1:500; Abcam. Cat # ab53554), Rabbit anti-PGP 9.5 (1:200; Abcam. Cat # ab27053), Rabbit anti-RET (1:100; Alomone labs, Israel, Cat #: ANT-025), goat anti-GFP (1:200; Rockland Cat # 600-101-215), Rabbit anti-S100b (1:100; Neomarkers, Cat # RB-044-A). Rabbit anti-Calretinin (Invitrogen 18-0211) Anti-Rabbit nNOS (Santa Cruz Cat# SC648), Anti-rabbit synaptophysin (Abcam Cat# ab1469). Secondary antibodies included goat anti-mouse IgG Alexa Fluor 546, goat anti-rabbit Alexa Fluor 488, donkey anti-goat Alexa Fluor 488, donkey anti-goat Alexa Fluor 546, donkey anti-mouse Alexa Fluor 546 and goat anti-human Alexa Fluor 594, all from Invitrogen (Carlsbad, CA). Cell nuclei were stained with DAPI (Vector Labs, Burlingame, CA).

Belkind-Gerson J., Graham H.K., Reynolds J., Hotta R., Nagy N., Cheng L., Kamionek M., Shi H.N., Aherne C.M, & Goldstein A.M. (2017). Colitis promotes neuronal differentiation of Sox2+ and PLP1+ enteric cells. Scientific Reports, 7, 2525.

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Immunofluorescence Staining Protocol

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Standard IF staining was performed according to the schematic shown in Figure 1a. First, sections were dewaxed in xylene and rehydrated in descending ethanol concentrations. Second, antigen retrieval was done by heating the sections to 95°C in 0.01 mol/L citric acid buffer (pH 6.0) for 15 min. Third, nonspecific sites were blocked by incubating the sections with normal goat serum in PBS for 30 min at 37°C. Fourth, the sections were incubated, respectively, with rabbit anti-PGP 9.5 (1:100 dilution, Abcam, ab8189, MA, USA), rabbit anti-TH (1:1000 dilution, Sigma, T8700, NJ, USA), or rabbit anti-VIP (1:1000 dilution, Immunostar, 20077, WI, USA), at 4°C overnight, followed by incubation with Alexa Fluor 488-labeled goat anti-rabbit secondary antibody (1:500, Beyotime, A0423, Jiangsu, China) for 1 h in the dark at room temperature. Finally, sections were counterstained with 5 μg/ml 4’, 6-diamidino-2-phenylindole (Beyotime, Jiangsu, China) for 10 min in the dark at room temperature and mounted with antifade mounting medium (Beyotime, Jiangsu, China). Sections were washed three times with PBS between steps. The results were observed with a fluorescence microscope (Olympus BX51, Tokyo, Japan).

Ouyang Z., Li H.H., Zhang M.J., Xie S.T, & Cheng L.H. (2018). Differential Innervation of Secretory Coils and Ducts in Human Eccrine Sweat Glands. Chinese Medical Journal, 131(16), 1964-1968.

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Immunohistochemical Analysis of Chronic Pain

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On 14 and 28 days after induction of EAP based on pelvic tactile allodynia response, mice were anesthetized with 2.5% isoflurane (Isothesia; Butler) and transcardially perfused with cold PBS followed by ice-cold 4% paraformaldehyde. DRGs, spinal cords of cervical, lumbar and sacral segments and prostates were removed and post-fixed overnight in 4% paraformaldehyde. Then the tissues were transferred to 30% sucrose solution overnight. Tissues were mounted with ornithine carbamyl transferase embedding medium (Tissue Tek) on dry ice and stored at −80°C for further experiments. The antibodies used in this study were as follows: rabbit anti-TMEM199 (1:1,000, Abcam), goat anti-IBA1 (1:1,000, Novus), rat anti-CD45 (1:500, Abcam), guinea pig anti-NeuN (1:1,000, Synaptic Systems), rabbit anti-PGP9.5 (1:500, Abcam), mouse anti-CCL2 (1:200, Proteintech) and chicken anti-GFAP (1:1,000, Novus). The multiple immunolabelings were visualized respectively by using Cy^tm2- or Cy^tm3-conjugated-donkey anti-rabbit IgG, Cy^tm3-conjugated-donkey anti-mouse IgG, Cy^tm3 or 5-conjugated-donkey anti-goat IgG, Cy^tm3-conjugated-donkey anti-rat IgG, Cy^tm5-conjugated-donkey anti-Guinea Pig IgG, Brilliant Violet 421-conjugated Donkey Anti-Chicken IgG (1:1,000, Jackson ImmunoResearch). Diamond Antifade mounting medium (Invitrogen) with or without DAPI (dependent on wavelength assignment in multicolor labeling).

Liu Z., Murphy S.F., Wong L., Schaeffer A.J, & Thumbikat P. (2020). Neuronal/astrocytic expression of chemokine (C-C motif) Ligand 2 is associated with monocyte/macrophage recruitment in male chronic pelvic pain. Pain, 161(11), 2581-2591.

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