Revision 1

#33738Store at -20C

1 Kit

(9 x 20 microliters)

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Product Includes Product # Quantity Mol. Wt Isotype/Source
Complexin-1 (D5Q5H) Rabbit mAb 17700 20 µl 14 kDa Rabbit IgG
Complexin-1/2 (D8A6E) Rabbit mAb 28070 20 µl 14-16 kDa Rabbit IgG
Munc18-1 (D4O6V) Rabbit mAb 13414 20 µl 70 kDa Rabbit IgG
SNAP25 (D7B4) Rabbit mAb 5308 20 µl 25 kDa Rabbit IgG
Synaptophysin (D8F6H) XP® Rabbit mAb 36406 20 µl 38 kDa Rabbit IgG
Synaptotagmin-1 (D33B7) Rabbit mAb 14558 20 µl 60 kDa Rabbit IgG
Synapsin-1 (D12G5) XP® Rabbit mAb 5297 20 µl 77 kDa Rabbit IgG
Syntaxin 1A (D4E2W) Rabbit mAb 18572 20 µl 33 kDa Rabbit IgG
VAMP2 (D6O1A) Rabbit mAb 13508 20 µl 13 kDa Rabbit 
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl Goat 

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

The Presynaptic Vesicle Cycle Antibody Sampler Kit provides an economical means of detecting expression of key synaptic vesicle proteins. The kit includes enough antibodies to perform two western blot experiments with each primary antibody.

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibodies.

Background

The synapse is the unit of information transfer in the brain between neurons (1). Synaptic transfer is initiated by action potential-initiated synaptic vesicles fusion at the presynaptic terminal to release neurotransmitters. The basic molecular machinery that drives all membrane fusion events in all cells include N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins that form a trans-ternary complex between to-be-fused membranes. The zippering up of SNARE proteins provides the energy required for membrane fusion. At the synapse, the SNARE proteins include 25 kDa synaptosome-associated protein (SNAP25), syntaxin 1A (STX1A), and vesicle-associated membrane protein 2 (VAMP2, also called synaptobrevin). Neurotransmitter release, however, is a highly regulated process that occurs in response to action potential-initiated Ca2+-influx. Regulation of synaptic vesicle fusion is mediated by SNARE-binding proteins like complexin and the Ca2+-associated vesicle protein, synaptotagmin-1, that cooperatively function to initiate Ca2+-induced synaptic vesicle fusion. The synaptic vesicle cycle represents a cycle of events that prime the synaptic vesicle before fusion and retrieve fused membrane after full fusion. Synaptic vesicle-associated proteins like synaptophysin and synapsin-1, as well as target membrane proteins like Munc18-1, play a complex role in priming synaptic vesicles to be fusion competent.   

  1. Südhof, T.C. and Rothman, J.E. (2009) Science 323, 474-7.

Background References

    Trademarks and Patents

    Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
    XP is a registered trademark of Cell Signaling Technology, Inc.
    All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

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    Revision 1
    #33738

    Presynaptic Vesicle Cycle Antibody Sampler Kit

    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 1 Expand Image
    Simple Western™ analysis of lysates (0.1 mg/mL) from rat brain using Munc18-1 (D4O6V) Rabbit mAb #13414. The virtual lane view (left) shows the target band (as indicated) at 1:50 and 1:250 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:50 (blue line) and 1:250 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 2 Expand Image
    Simple Western™ analysis of lysates (0.1 mg/mL) from mouse brain using Synaptophysin (D8F6H) XP® Rabbit mAb #36406. The virtual lane view (left) shows the target band (as indicated) at 1:10 and 1:50 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 3 Expand Image
    Simple Western™ analysis of lysates (0.1 mg/mL) from CAD cells using Synapsin-1 (D12G5) XP Rabbit mAb #5297. The virtual lane view (left) shows the target band (as indicated) at 1:10 and 1:50 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 4 Expand Image
    Western blot analysis of extracts from mouse brain, rat brain, and human cortex using Munc18-1 (D4O6V) Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 5 Expand Image
    Western blot analysis of extracts from various cell lines and tissues using VAMP2 (D6O1A) Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 6 Expand Image
    Western blot analysis of extracts from mouse and rat brain using Synaptotagmin-1 (D33B7) Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 7 Expand Image
    Western blot analysis of extracts from various cells lines and tissues using Complexin-1 (D5Q5H) Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 8 Expand Image
    Western blot analysis of extracts from mouse brain, mouse lung, and rat brain using Syntaxin 1A (D4E2W) Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 9 Expand Image
    Western blot analysis of extracts from Neuro-2a, NIH/3T3, and mouse brain, using Complexin-1/2 (D8A6E) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 10 Expand Image
    Western blot analysis of extracts from mouse and rat tissue using Synaptophysin (D8F6H) XP® Rabbit mAb (upper) and B-Actin (D6A8) Rabbit mAb #8457 (lower).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 11 Expand Image
    Western blot analysis of extracts from CAD cells and neonatal mouse brain using Synapsin-1 (D12G5) XP® Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 12 Expand Image
    Western blot analysis of extracts from mouse and rat brain using SNAP25 (D7B4) Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 13 Expand Image
    After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 14 Expand Image
    Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing Myc-tagged full-length human VAMP1 (hVAMP1-Myc; +) or VAMP2 (hVAMP2-Myc; +), using VAMP2 (D6O1A) Rabbit mAb (upper) and Myc-Tag (71D10) Rabbit mAb #2278 (lower).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 15 Expand Image
    Confocal immunofluorescent analysis of mouse retina using Synaptotagmin-1 (D33B7) Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 16 Expand Image
    Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing Myc-tagged full length human complexin-1 (hCPLX-1-Myc; +) or complexin-2 (hCPLX-2-Myc; +), using Complexin-1 (D5Q5H) Rabbit mAb (upper), and Myc-Tag (71D10) Rabbit mAb #2278 (lower).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 17 Expand Image
    Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing Myc-tagged full-length human complexin-1 (hCPLX-1-Myc; +) or complexin-2 (hCPLX-2-Myc; +), using Complexin-1/2 (D8A6E) Rabbit mAb (upper), and Myc-Tag (71D10) Rabbit mAb #2278 (lower).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 18 Expand Image
    Immunohistochemical analysis of paraffin-embedded Neuro-2a cell pellet (left, positive) or C2C12 cell pellet (right, negative) using Synaptophysin (D8F6H) XP® Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 19 Expand Image
    Immunohistochemical analysis of paraffin-embedded human cerebellum using Synapsin-1 (D12G5) XP® Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 20 Expand Image
    Immunoprecipitation of VAMP2 from Neuro-2a cell extracts using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or VAMP2 (D6O1A) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using VAMP2 (D6O1A) Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 21 Expand Image
    Simple Western™ analysis of lysates (0.5 mg/mL) from IMR-32 cells using Synaptotagmin-1 (D33B7) Rabbit mAb #14558. The virtual lane view (left) shows the target band (as indicated) at 1:10 and 1:50 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 22 Expand Image
    Immunoprecipitation of Complexin-1 from Neuro-2a cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 and lane 3 is Complexin-1 (D5Q5H) Rabbit mAb. Western blot was performed using Complexin-1 (D5Q5H) Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 23 Expand Image
    Immunoprecipitation of complexin-1/2 from Neuro-2A cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Complexin-1/2 (D8A6E) Rabbit mAb. Western blot analysis was performed using Complexin-1/2 (D8A6E) Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 24 Expand Image
    Immunohistochemical analysis of paraffin-embedded mouse cerebellum using Synaptophysin (D8F6H) XP® Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 25 Expand Image
    Immunohistochemical analysis of paraffin-embedded mouse cortex using Synapsin-1 (D12G5) XP® Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 26 Expand Image
    Immunohistochemical analysis of paraffin-embedded human colon using VAMP2 (D6O1A) Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 27 Expand Image
    Confocal immunofluorescent analysis of Neuro-2a (positive, left panels) and NIH/3T3 (negative, right panels) using Complexin-1 (D5Q5H) Rabbit mAb (green). Actin filaments were labeled with DyLight 554 Phalloidin #13054 (red, upper panels). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 28 Expand Image
    Confocal immunofluorescent analysis of Neuro-2a (left) and NIH/3T3 (right) cells, using Complexin-1/2 (D8A6E) Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 29 Expand Image
    Immunohistochemical analysis of paraffin-embedded mouse colon using Synaptophysin (D8F6H) XP® Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 30 Expand Image
    Immunohistochemical analysis of paraffin-embedded rat retina using Synapsin-1 (D12G5) XP® Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 31 Expand Image
    Immunohistochemical analysis of paraffin-embedded rat kidney using VAMP2 (D6O1A) Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 32 Expand Image
    Immunohistochemical analysis of paraffin-embedded human non-small cell lung carcinoma using Synaptophysin (D8F6H) XP® Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 33 Expand Image
    Confocal immunofluorescent analysis of mouse brain using Synapsin-1 (D12G5) XP® Rabbit mAb (green) and β3-Tubulin (TU-20) Mouse mAb #4466 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 34 Expand Image
    Immunohistochemical analysis of paraffin-embedded mouse pancreas using VAMP2 (D6O1A) Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 35 Expand Image
    Immunohistochemical analysis of paraffin-embedded human endometrioid adenocarcinoma using Synaptophysin (D8F6H) XP® Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 36 Expand Image
    Confocal immunofluorescent analysis of normal rat retina using VAMP2 (D6O1A) Rabbit mAb (green) and GFAP (GA5) Mouse mAb (Alexa Fluor® 555 Conjugate) #3656 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 37 Expand Image
    Immunohistochemical analysis of paraffin-embedded mouse pancreas using Synaptophysin (D8F6H) XP® Rabbit mAb.
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 38 Expand Image
    Confocal immunofluorescent analysis of PC-12 cells using VAMP2 (D6O1A) Rabbit mAb (green). Actin filaments were labeled with DyLight 554 Phalloidin #13054 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 39 Expand Image
    Confocal immunofluorescent analysis of mouse retina using Synaptophysin (D8F6H) XP® Rabbit mAb (green), Neurofilament-L (DA2) Mouse mAb #2835 (yellow) and Lamin A/C (4C11) Mouse mAb #4777 (red). Samples were mounted in ProLong® Gold Antifade Reagent with DAPI #8961 (blue).
    Presynaptic Vesicle Cycle Antibody Sampler Kit: Image 40 Expand Image
    Confocal immunofluorescent analysis of mouse brain using Synaptophysin (D8F6H) XP® Rabbit mAb (green), Neurofilament-L (DA2) Mouse mAb #2835 (yellow) and Lamin A/C (4C11) Mouse #4777 (red). Samples were mounted in ProLong® Gold Antifade Reagent with DAPI #8961 (blue).