MA5-14532

antibody from Invitrogen Antibodies

Targeting: SYP MRX96

Provider product page for MA5-14532

Western blot

Antibody data

Antibody Data
Antigen structure
References [49]
Comments [0]
Validations
Western blot [3]
Immunocytochemistry [1]
Immunohistochemistry [3]
Flow cytometry [1]
Other assay [8]

Submit

Product number: MA5-14532 - Provider product page
Provider: Invitrogen Antibodies
Product name: Synaptophysin Monoclonal Antibody (SP11)
Antibody type: Monoclonal
Antigen: Synthetic peptide
Description: MA5-14532 targets Synaptophysin in IHC (P) and WB applications and shows reactivity with Human samples. The MA5-14532 immunogen is a synthetic peptide of human synaptophysin.
Reactivity: Human, Mouse, Rat
Host: Rabbit
Isotype: IgG
Antibody clone number: SP11
Vial size: 500 µL
Concentration: Conc. Not Determined
Storage: Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

SYP protein structure - MA5-14532 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot of Synaptophysin using Synaptophysin Monoclonal Antibody (Product # MA5-14532) on IMR-5 Cells.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on tissue, whole cell extracts (30 µg lysate) of Rat Brain (Lane 1), Mouse Brain (lane 2), and SH-SY5Y (lane 3).The blots were probed with Anti-Synaptophysin Rabbit Monoclonal Antibody (Product # MA5-14532, 1:1000) and detected by chemiluminescence Goat Anti-Rabbit IgG Secondary Antibody, HRP conjugate (Product # G-21234, 1:5000 dilution). A 34 kDa band corresponding to Synaptophysin was observed across cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 10 % Bis-Tris gel (Product # NP0301BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5 % skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockout of SYP was achieved by CRISPR-Cas9 genome editing using LentiArray™ Lentiviral sgRNA (Product # A32042) (Assay ID CRISPR758834_LV) and LentiArray Cas9 Lentivirus (Product # A32064). Western blot analysis of SYP was performed by loading 30 µg of SH-SY5Y wild type (Lane 1), SH-SY5Y CAS9 (Lane 2), SH-SY5Y SYP KO (Lane 3) whole cell extracts. The blot was probed with Anti-Synaptophysin Monoclonal Antibody (SP11)(Product # MA5-14532) using 1:1000 dilution and Goat anti-Rabbit IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036). Loss of signal upon CRISPR mediated knockout (KO) using the LentiArray™ CRISPR product line confirms that antibody is specific to SYP.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Formalin-fixed, paraffin-embedded neuroendocrine tumor stained with Synaptophysin using peroxidase-conjugate and AEC. Note cytoplasmic staining of tumor cells.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of Synaptophysin showing staining in the cytoplasm/synaptophysin of paraffin-embedded human cerebellum tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a Synaptophysin Rabbit Monoclonal Antibody (Product # MA5-14532) diluted in 3% BSA-PBS at a dilution of 1:20 for 1 hour at 37ºC in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of Synaptophysin showing staining in the cytoplasm/synaptophysin of paraffin-embedded mouse brain tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a Synaptophysin Rabbit Monoclonal Antibody (Product # MA5-14532) diluted in 3% BSA-PBS at a dilution of 1:20 for 1 hour at 37ºC in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: 5 Figure The effect of CM on synaptic markers in the peri-ischemic cortex. Representative micrographs of (a) GAP-43 and (b) SYP immunohistochemical staining in peri ischemic rat brains. Scale bar = 100 mum. ( n = 3). The protein levels of (c) GAP-43, (d) SYP, (e) CREB, and (f) p-CREB/CREB ratio significantly decreased in the peri-ischemic cortex on Day 7 following MCAO. Three ICV injections of CM could restore levels of GAP-43, SYP, and p-CREB up to the normal levels observed in the sham group. Data are reported as mean +- SEM ( n = 4). ** p < .01 and *** p < .001 versus Sham, ### p < .001 versus MCAO + NS, +++ p < .001 versus CM1. CM, conditioned medium; CREB, cAMP response element-binding protein; DMEM, Dulbecco's modified Eagle's medium; ICV, intracerebroventricular; GAP-43, growth-associated protein-43; MCAO, middle cerebral artery occlusion; NS, normal saline; SEM, standard error of mean; SYP, synaptophysin

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: 335.g003 Fig 3 Functional synapses between cells. A. Synapses between neurons (A1/N4); B. Synapses between neurons and astrocytes (A1/N4); C. No synapses in A4/N1 group. D. 3D co-culture with synapses (A1/N4). Red: Synaptophysin for synapses, green: beta-tubulin III for neurons, white: GFAP for astrocytes, and blue: Hoechst for nuclei.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 3 SCLC neuroendocrine subtypes segregate by ferroptosis response. a , b Eight human SCLC cell lines were treated as indicated for 24 h, cell viability was determined by Cell Titer Blue. c The indicated human SCLC cells were treated with DMSO, RSL3 [100 nM] or RSL3/Fer-1 [5 uM] for 5 h and stained for lipid ROS accumulation using BODIPY C11. Cells were analyzed by flow cytometry. Gray = DMSO treated, violet = RSL3 treated, turquoise = RSL3/Fer1 treated. d RNA-seq data of human SCLC lines (sensitive n = 3 cell lines, H524, NCIH82, H526; resistant n = 5 cell lines, NCIH889, NCIH1092, CORL88, H1836, H2171) were analyzed for differential expression between responders and non-responders, heatmap represents hierarchical clustering of FPKM (log2 + 0.01) of the 50 most differentially expressed genes. Heatmap color code indicates expression levels between each sample and the average of each gene, dendrogram shows the distance between sample populations. e ASCL1 and REST1 expression (FPKM (log2 + 0.01) comparing three sensitive and five resistant cell lines is plotted, boxplot center line, mean; box limits, upper and lower quartile; whiskers min. to max. f Gene set enrichment analysis (GSEA) of a ranked list from ferroptosis sensitive and resistant cells was performed. g Western blot of SCLC NE subtype marker expression in the indicated manually separated stickers and floaters lines ( n = 3). h Manually separated stickers and floaters lines ( n = 3) were treated with RSL3 [1 uM] fo

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 4 NE induces NED markers in PC cells. A Immunostaining shows that 50 uM NE induces the expression of the NED markers CHGB and SYP in DU145 cells at 24 h. Both CHGB and SYP show characteristic granular staining after NE treatment. B 50 uM NE induces the expression of CHGB and SYP in LNCaP cells at 24 h. The control cells display weak basal expression of CHGB and SYP, but NE treatment upregulates the expression of these markers, evident by their increased staining intensity. Scale bar, 20 um.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: 1 FIGURE alpha-Synuclein preformed fibrils (PFF) preparation, Characterization, and SH-SY5Y neuroblastoma cell differentiation. (A) Characterization of purified alpha-syn fractions on 15% SDS-PAGE eluted with different NaCl concentrations. (B) Western blot of purified alpha-syn. Western blot analysis of purified alpha-syn, primary antibody: H3C And secondary: Anti-mouse HRP conjugated antibody. (C) Thioflavin-T assay of alpha-synuclein (Red Dots) aggregation. The data represent +-SEM of three independent replicates. (D) Representative fluorescence microscopy image of alpha-synuclein fibrils. Scale bar indicates 5 mum: (E) Representative AFM image of alpha-synuclein fibrils. The scale bar indicates 500 nm. (F-K) show the confocal microscopy images represented as maximum z projection (F-G), Phalloidin (F-Actin) staining in undifferentiated SH-SY5Y cells at day 0, and differentiated neurons on day 21, respectively. (I and J) FM 1-43 dye staining in undifferentiated SH-SY5Y cells at day 0 and differentiated neurons on day 21, respectively. The cells were stained with FM 1-43FX (5 mug/ml) dye for 15 min at 37degC, followed by cell fixation with 4% PFA. (H and K) shows the immunofluorescence analysis done on differentiated neurons on day 21 for Tau, Synaptophysin (SP11) and Microtubule associated protein (MAP2). Mature neurons confirmation was done against Tau, SP11 and MAP2 at 1:250 dilution for 3 h followed by secondary antibody labeling with Goat anti-Rabbit IgG (H + L) Highly C

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 1 MYCN Drives Transformation of NES Cells to Medulloblastoma (A) Schematic showing differentiation of iPSCs to NES cells, transduction with FLAG-MYCN, and orthotopic implantation into mice. (B) Western blot showing misexpressed MYCN in normal WTC10 NES cells. (C) Empty vector and MYCN NES cells were treated with 5-ethynyl-2'-deoxyuridine (EdU) for 1 h and analyzed via flow cytometry. MYCN NES cells show increased EdU incorporation and S-phase fraction. Data are presented as mean +- SEM; * p < 0.05 (t test). (D) CyQuant Direct cell proliferation analysis showing increased proliferation in MYCN NES cells. Data are presented as mean +- SEM; * p < 0.05 (t test). (E) Kaplan-Meier survival curve of mice injected with empty vector and MYCN NES cells (n = 4). p = 0.004 (log-rank test). (F) (Left) Low magnification (50x) of H&E staining of WTC10 MYCN tumors showing implanted MYCN NES cells expanding and distorting the mouse cerebellum and invading down Virchow-Robin spaces (arrow). (Middle) High magnification (400x) of H&E staining revealing anaplastic features including frequent mitoses (arrows), cell-cell wrapping (black arrowhead), prominent nuclei (red arrowhead), and karyorrhexis (red arrow) characteristic of large cell and/or anaplastic medulloblastoma. (Right) Immunohistochemical staining for synaptophysin (SYP) highlighting neuroblastic pseudorosettes. Scale bars represent 200 mum for 50x images and 20 mum for 400x images. See also Figures S1 and S2 and Tables S1 , S4 ,

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 2 DeltaGluN1 induced loss of dendritic spines in CA1 neurons. (A) Representative confocal images demonstrating co-localization of synaptophysin and DIL neurotracer in the CA1 (pyr, pyramidal layer; mol, molecular layer). Scale bar = 20 mum and 5 mum. (B) Bar graph [One-Way analysis of variance (ANOVA)] illustrating statistical comparison of CA1 dendritic spine count (DIL/synaptophysin). (C) Bar graph (One-Way ANOVA) comparing cytoskeletal filament count in hippocampal dendritic spines. (D) Transmission electron microscopy (TEM) photomicrographs illustrating hippocampal synapses. Yellow arrowheads indicate the synaptic cytoskeleton. Scale bar = 1 mum. (E) TEM photomicrographs demonstrating post-synaptic densities (PSD) in the hippocampus. Scale bar = 1 mum. (F) Bar graph illustrating the comparative thickness of PSDs ( B,C,F ; ** p < 0.01, *** p < 0.001).