anti ib4 fitc  (Vector Laboratories)

Journal: bioRxiv

Article Title: Isolectin B4 (IB4)-conjugated streptavidin for the selective knockdown of proteins in IB4-positive (+) nociceptors

doi: 10.1101/2023.12.18.572242

Figure Lengend Snippet: Knockdown of PKCε in IB4(+) C-fiber afferents. Representative images from L4/L5 DRG from rats treated with either IB4-streptavidin biotinylated PKCε sense ODN (upper image panels) or IB4-streptavidin biotinylated PKCε antisense ODN (lower image panels). Color coding: Red = PKCε immunoreactivity, Blue = DAPI, Green = IB4 immunoreactivity, Yellow = PKCε/IB4 colocalization. Note that up to 40% of all DRG neurons in the rat are IB4 positive (+) and the vast majority are small diameter C-fiber DRG neurons. PKCε immunoreactivity can be observed in small-, medium-, and large-diameter DRG neurons. Interestingly, the vast majority of PKCε expressing DRG (∼90%) are IB4 positive (+). Scale bar = 50 µm.

Article Snippet: Sections were incubated overnight at 4°C with a 1:500 dilution of a Rabbit (Rb) anti-PKCε antibody (sc-214, Santa Cruz Biotechnology) in antibody dilution buffer, washed three times for 10 minutes each at RT with TBST and probed for 2h at RT with a 1:500 dilution of an Alexa Fluor conjugated Goat anti-Rb antibody (Invitrogen cat. #: A11012) and a 1:1000 dilution of IB4-FITC (Invitrogen, cat. #: I21411) in antibody dilution buffer supplemented with 0.1mM CaCl 2 , 0.1mM MnCl 2 , 0.1mM MgCl 2 for 2h at RT ( ).

Techniques: Expressing

Journal: CNS Neuroscience & Therapeutics

Article Title: The G protein‐coupled estrogen receptor of the trigeminal ganglion regulates acute and chronic itch in mice

doi: 10.1111/cns.14367

Figure Lengend Snippet: Expression and distribution of the G protein‐coupled estrogen receptor (GPER) in trigeminal ganglion (TG) neurons of the mouse. (A) Schematic diagram of cholera toxin subunit B (CTB) injection in mouse and representative immunofluorescence images of CTB‐labeled TG neurons innervating the mouse cheek colocalized with GPER, along with the quantitative analysis of the overlap percentage. (B–D) Double immunofluorescence staining showing the colocalization of GPER with IB4 (B), CGRP (C) and NF200 (D), along with quantitative analysis of the overlap percentage ( n = 4 mice per group, scale bar: 100 or 40 μm).

Article Snippet: Primary antibodies used in these experiments were rabbit anti‐GPER (1:1000, Lifespan), rabbit anti‐c‐Fos (1:500, Abcam), mouse anti‐neuroflament‐200 (NF200; 1:1000, Abcam), mouse anti‐calcitonin gene‐related peptide (CGRP; 1:100, Abcam), mouse anti‐isolectin B4 conjugated to fluorescein isothiocyanate (IB4‐FITC; 1:1000, Sigma), rabbit anti‐TRPA1 (1:500, Abcam), and guinea pig anti‐TRPV1 (1:1000, Millipore).

Techniques: Expressing, Injection, Immunofluorescence, Labeling, Double Immunofluorescence Staining

Journal: CNS Neuroscience & Therapeutics

Article Title: The G protein‐coupled estrogen receptor of the trigeminal ganglion regulates acute and chronic itch in mice

doi: 10.1111/cns.14367

Figure Lengend Snippet: Gper deficiency enhances the function of TRPA1 and TRPV1 in TG neurons. (A) Representative immunofluorescence images showing the colocation of GPER with TRPA1. (B) Quantitative analysis of the percentage of GPER + /TRPA1 + neurons in total GPER + neurons of the TG ( n = 4 mice per group, scale bar: 100 or 40 μm). (C) Representative immunofluorescence images showing colocation of GPER and TRPV1. (D) Quantitative analysis of the percentage of GPER + /TRPV1 + neurons in total GPER + neurons of the TG ( n = 4 mice per group, scale bar: 100 or 40 μm). (E) Representative fluorescence images of mouse TG neurons loaded with Fura‐2 at the baseline and after application of allyl isothiocyanate (AITC, 100 μM) or capsaicin (Cap, 1 μM). Arrows indicate TG neurons responsive to the agent (scale bar: 200 μm). (F) Representative traces of Ca 2+ responses evoked by AITC or Cap in TG neurons of WT (black) and Gper −/− (red) mice. Black bars above the traces show the duration of the chemical treatment. (G–J) In comparison with WT mice, the percentage of AITC‐responsive (G) or Cap‐responsive neurons (I) was significantly increased in Gper −/− mice (** p < 0.01, *** p < 0.001, χ 2 test). In comparison with WT mice, the Ca 2+ signal amplitude induced by AITC (H) or Cap (J) was significantly higher in Gper −/− mice (** p < 0.01, *** p < 0.001, unpaired Student's t‐ test).

Article Snippet: Primary antibodies used in these experiments were rabbit anti‐GPER (1:1000, Lifespan), rabbit anti‐c‐Fos (1:500, Abcam), mouse anti‐neuroflament‐200 (NF200; 1:1000, Abcam), mouse anti‐calcitonin gene‐related peptide (CGRP; 1:100, Abcam), mouse anti‐isolectin B4 conjugated to fluorescein isothiocyanate (IB4‐FITC; 1:1000, Sigma), rabbit anti‐TRPA1 (1:500, Abcam), and guinea pig anti‐TRPV1 (1:1000, Millipore).

Techniques: Immunofluorescence, Fluorescence, Comparison

Journal: Frontiers in Neurology

Article Title: Attenuation of immobilization stress-induced hypertension by temperature-controllable warm needle acupuncture in rats and the peripheral neural mechanisms

doi: 10.3389/fneur.2023.1168012

Figure Lengend Snippet: Immunohistochemistry for TRPV1 and/or CGRP, NF200 and IB4 in dorsal root ganglia (DRGs) following warm needle acupuncture instrument (WAI) stimulation at PC-6. (A,B) Fluorescence intensity and representative images of TRPV1 and CGRP double-stained DRG neurons (yellow) in the normal ( n = 6) and WAI ( n = 6) groups. *** indicates a significance level of p < 0.001 vs. Normal. Immunofluorescent images for TRPV1 (red) and CGRP (green) (C,D) Fluorescence intensity and representative images of TRPV1- and NF200-positive DRG neurons in the normal ( n = 6) and WAI ( n = 6) groups. Immunofluorescent images for TRPV1 (red) and NF200 (green) (E,F) Fluorescence intensity and representative images of TRPV1- and IB4-positive DRG neurons in the normal ( n = 6) and WAI ( n = 6) groups. * p < 0.05 vs. Normal. Double-stained cells (yellow) by TRPV1 (green)/IB4 (red). Scale bar: 100 μm.

Article Snippet: The slides were incubated with mouse anti-TRPV1 antibody (1:500, Santa Cruz Biotechnology), rabbit anti-calcitonin gene-related peptide (CGRP, 1:1000, Abcam), anti-lectin IB4 Alexa Fluor 488 conjugated to FITC (1:500, Invitrogen), rabbit anti-neurofilament 200 (NF200, 1:1000, Sigma–Aldrich) and mouse monoclonal PGP 9.5 antibody (1:1000; Abcam, Cambridge, United Kingdom; ab8189) overnight at 4°C, followed by incubation with secondary antibodies (1:200, Alexa Fluor 488 donkey anti-rabbit IgG antibody), Thermo Scientific, MA, United States, Product# A-21206; 1:200, Alexa Fluor ® 594 donkey anti-mouse IgG antibody, Thermo Scientific, Product# A-21203, donkey anti-mouse IgG Cy3 (1:500, 133,644, Invitrogen).

Techniques: Immunohistochemistry, Fluorescence, Staining