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CellPaint™ TSP membrane stain

HeLa cells were labeled with CellPaint™ TSP membrane stain (Cat No. 22700). Labeled cells were imaged on the Keyence BZ-X710 All-In-One Fluorescence Microscope equipped with a Cy3/TRITC filter set.
HeLa cells were labeled with CellPaint™ TSP membrane stain (Cat No. 22700). Labeled cells were imaged on the Keyence BZ-X710 All-In-One Fluorescence Microscope equipped with a Cy3/TRITC filter set.
HeLa cells were labeled with CellPaint™ TSP membrane stain (Cat No. 22700). Labeled cells were imaged on the Keyence BZ-X710 All-In-One Fluorescence Microscope equipped with a Cy3/TRITC filter set.
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Physical properties
Molecular weight1016.27
SolventDMSO
Spectral properties
Excitation (nm)496
Emission (nm)633
Storage, safety and handling
Intended useResearch Use Only (RUO)
StorageFreeze (< -15 °C); Minimize light exposure

OverviewpdfSDSpdfProtocol


See also: Plasma Membrane
Molecular weight
1016.27
Excitation (nm)
496
Emission (nm)
633
TSP is a styrylpyridine-based fluorescent membrane probe suitable for imaging plasma membranes in living cells and tissues. It was reported by Guo et al in 2016 (Analyst, 2016, 141, 3228). The probe is a molecular rotor that has fluorescence sharply enhanced in viscous media. Its fluorescence is also microenvironment-sensitive, enables the turn-on imaging of plasma membranes with a high signal-to-noise ratio. Guo et al has demonstrated that TSP has high photostability, low cytotoxicity and excellent biocompatibility. It can also be used in 2 photon imaging.

Platform


Fluorescence microscope

ExcitationCy3/TRTC filter set
EmissionCy3/TRITC filter set
Recommended plateBlack wall/clear bottom

Example protocol


AT A GLANCE

  1. Prepare cells in growth medium
  2. Incubate cells with Cellpaint™ TSP membrane stain
  3. Analyze under fluorescence microscope with Cy3/TRITC filter set 

Chemical and Physical Properties
Molecular Weight: 1016.27
Solvent: dimethyl sulfoxide (DMSO)
Spectral Properties: Excitation = 490 nm; Emission = 600 nm

CELL PREPARATION

For guidelines on cell sample preparation, please visit https://www.aatbio.com/resources/guides/cell-sample-preparation.html

PREPARATION OF STOCK SOLUTIONS

Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.

Cellpaint™ TSP membrane stain stock solution (500X)
  1. Thaw the vial of CellPaint™ TSP membrane stain at room temperature before use.
  2. Add 100 µL of DMSO into the vial of Cellpaint™ TSP membrane stain to make 500X stock solution.
    Note     Unused Cellpaint™ TSP membrane stain stock solution can be aliquoted and stored at ≤ -20 °C for a couple of months if the tubes are sealed tightly. Protect from light and avoid repeated freeze-thaw cycles. 

PREPARATION OF WORKING SOLUTION

Cellpaint™ TSP membrane stain working solution (10X)
  1. Add 20 µL of 500X stock solution into 1 mL of cell culture medium, and mix well.
    Note     We recommend making the working solution fresh before use.
    Note     20 µL of 500X Cellpaint™ TSP membrane stain stock solution is enough for one 96-well plate.
    Note     We recommend using a cell culture medium to make a working solution for better resolution images. 

SAMPLE EXPERIMENTAL PROTOCOL

Stain Cells
  1. Add 10 µL/well (96-well plate/100 µL volume) or 5 µL/well (384-well plate-50 µL volume) of 10X Cellpaint™ TSP membrane stain working solution in the cell plate. Incubate the cells at 37 °C for 30-60 minutes, protected from light.
    Note     The optimal concentration of the cell membrane probe varies depending on the specific application. The staining conditions may be modified according to the particular cell type and the permeability of the cells or tissues to the probe.
  2. Remove working solution in each well. Wash cells with physiological buffer (such as HHBS, DPBS or buffer of your choice) for three times and replace with HHBS.
  3. Observe the fluorescence signal in cells using fluorescence microscope with a Cy3/TRITC filter set. 

Calculators


Common stock solution preparation

Table 1. Volume of DMSO needed to reconstitute specific mass of CellPaint™ TSP membrane stain to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.

0.1 mg0.5 mg1 mg5 mg10 mg
1 mM98.399 µL491.995 µL983.99 µL4.92 mL9.84 mL
5 mM19.68 µL98.399 µL196.798 µL983.99 µL1.968 mL
10 mM9.84 µL49.2 µL98.399 µL491.995 µL983.99 µL

Molarity calculator

Enter any two values (mass, volume, concentration) to calculate the third.

Mass (Calculate)Molecular weightVolume (Calculate)Concentration (Calculate)Moles
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Spectrum


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spectrum

Spectral properties

Excitation (nm)496
Emission (nm)633

Images


Citations


View all 31 citations: Citation Explorer
Membrane trafficking and exocytosis are upregulated in port wine stain blood vessels
Authors: Yin, R., Rice, S. J., Wang, J., Gao, L., Tsai, J., Anvari, R. T., Zhou, F., Liu, X., Wang, G., Tang, Y., Mihm, M. C., Jr., Belani, C. P., Chen, D. B., Nelson, J. S., Tan, W.
Journal: Histol Histopathol (2019): 479-490
Cu(2+)-Directed Liposome Membrane Fusion, Positive-Stain Electron Microscopy, and Oxidation
Authors: Liu, Y., Liu, J.
Journal: Langmuir (2018): 7545-7553
Extraction of DNA from Sperm Cells in Mixed Stain by Nylon Membrane Bushing Separation Technique
Authors: Ma, J., Tong, Q., Gao, L. B., Zhu, C., Jiang, Z. Q.
Journal: Fa Yi Xue Za Zhi (2018): 417-419
Gram&apos;s Stain Does Not Cross the Bacterial Cytoplasmic Membrane
Authors: Wilhelm, M. J., Sheffield, J. B., Sharifian Gh, M., Wu, Y., Spahr, C., Gonella, G., Xu, B., Dai, H. L.
Journal: ACS Chem Biol (2015): 1711-7
The use of SMALPs as a novel membrane protein scaffold for structure study by negative stain electron microscopy
Authors: Postis, V., Rawson, S., Mitchell, J. K., Lee, S. C., Parslow, R. A., Dafforn, T. R., Baldwin, S. A., Muench, S. P.
Journal: Biochim Biophys Acta (2015): 496-501
Overview of electron crystallography of membrane proteins: crystallization and screening strategies using negative stain electron microscopy
Authors: Nannenga, B. L., Iadanza, M. G., Vollmar, B. S., Gonen, T.
Journal: Curr Protoc Protein Sci (2013): Unit17 15
Negative-stain electron microscopy of inside-out FtsZ rings reconstituted on artificial membrane tubules show ribbons of protofilaments
Authors: Milam, S. L., Osawa, M., Erickson, H. P.
Journal: Biophys J (2012): 59-68
Ultrastructure and retinal imaging of internal limiting membrane: a clinicopathologic correlation of trypan blue stain in macular hole surgery
Authors: Mackenzie, S. E., G and orfer, A., Rohleder, M., Schumann, R., Schlottmann, P. G., Bunce, C., Xing, W., Gregor, Z., Charteris, D. G.
Journal: Retina (2010): 655-61
A nitrobenzofuran-conjugated phosphatidylcholine (C12-NBD-PC) as a stain for membrane lamellae for both microscopic imaging and spectrofluorimetry
Authors: Hope-Roberts, M., Wainwright, M., Horobin, R. W.
Journal: Biotech Histochem (2008): 25-8
Indocyanine green-assisted internal limiting membrane peeling for macular holes to stain or not to stain?
Authors: Da Mata, A. P., Riemann, C. D., Nehemy, M. B., Foster, R. E., Petersen, M. R., Burk, S. E.
Journal: Retina (2005): 395-404