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Cell Meter™ Cell Adhesion Assay Kit

Cell adhesion measured with Cell Meter™ Cell Adhesion Assay Kit using a fluorescence microplate reader. Jurkat cells at different confluences or confluency levels were incubated in wells coated with different materials, and then stained with Calcein Ultragreen AM at 37°C for 30 mins. The signal was monitored at Ex/Em = 490/525 nm.
Cell adhesion measured with Cell Meter™ Cell Adhesion Assay Kit using a fluorescence microplate reader. Jurkat cells at different confluences or confluency levels were incubated in wells coated with different materials, and then stained with Calcein Ultragreen AM at 37°C for 30 mins. The signal was monitored at Ex/Em = 490/525 nm.
Ordering information
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Catalog Number23010
Unit Size
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Additional ordering information
Telephone1-408-733-1055
Fax1-408-733-1304
Emailsales@aatbio.com
InternationalSee distributors
ShippingStandard overnight for United States, inquire for international
Spectral properties
Excitation (nm)494
Emission (nm)514
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
UNSPSC12171501
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OverviewpdfSDSpdfProtocol


Excitation (nm)
494
Emission (nm)
514
The Cell Meter™ Cell Adhesion Assay Kit is a fast and sensitive assay for measuring cell-cell or cell-surface adhesion for a variety of cell types. In this assay, cells are labeled with Calcein UltraGreen AM and allowed to adhere. After removal of nonadherent cells, The fluorescence of Calcein UltraGreen is used to calculate the number of adherent cells. The use of our outstanding fluorogenic dye, Calcein UltraGreen AM provides a fast and sensitive method for measuring cell adhesion with a variety of cell types. Calcein UltraGreen AM is nonfluorescent but, once loaded into cells, is cleaved by endogenous esterases to produce highly fluorescent Calcein UltraGreen, a brightly fluorescent, pH-independent, cytoplasmic cell marker with the minimal interference to cell adhesion process. The Cell Meter™ cell adhesion assay is designed for use with fluorescence microplate readers. The robust performance of Calcein UltraGreen AM and simple procedure of the kit avoids problems associated with assays that utilize radioisotopes, which generate hazardous waste, and with assays that rely on the use of covalently coupled cell-surface labels, which can potentially alter cell function.

Platform


Fluorescence microscope

ExcitationFITC filter set
EmissionFITC filter set
Recommended plateBlack wall/clear bottom

Fluorescence microplate reader

Excitation490 nm
Emission525 nm
Cutoff515 nm
Recommended plateBlack wall/Clear bottom

Components


Component A: Calcein UltraGreen™ AM1 vial
Component B: Adhesion Assay Buffer1 bottle (25 mL)
Component C: DMSO1 vial (100 µL)

Example protocol


AT A GLANCE

Protocol summary
  1. Add cells on a plate coated with desired coating material
  2. Incubate cells at 37 °C to allow them to attach
  3. Remove the unattached cells
  4. Add Calcein Ultragreen AM working solution
  5. Incubate the cells at 37 °C for 20-30 minutes
  6. Remove supernatant and wash cells with HHBS or DPBS
  7. Measure the fluorescence intensity using fluorescence microplate reader with Ex/Em = 490/525 nm 

Important
Thaw all the kit components at room temperature before starting the experiment.

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.

Calcein Ultragreen AM stock solution
Add 50 µL of DMSO (Component C) into Calcein Ultragreen AM (Component A) and mix well.
Note     Store the unused Calcein Ultragreen AM stock solution at -20 °C in single use aliquots to avoid freeze thaw cycles.

PREPARATION OF WORKING SOLUTION

Calcein Ultragreen AM working solution
Add 50 µL of Calcein Ultragreen AM stock solution into 10 mL of Adhesion Assay Buffer and mix well.
Note     Calcein Ultragreen AM working solution should not be stored and should be used promptly.
Note     10 mL Calcein Ultragreen AM working solution is enough for 100 tests.

SAMPLE EXPERIMENTAL PROTOCOL

The following protocol can be used as a guideline and should be optimized according to the needs.
  1. Add 100 µL volumes of cells on a plate coated with desired coating material.
  2. Incubate plate at 37 °C for 2 to 3 hours.
    Note     For each cell line, optimal incubation time should be tested experimentally.
  3. Remove the growth medium and unattached cells.
  4. Add 100 µL of Calcein Ultragreen AM working solution and incubate plate at 37 °C for 20-30 minutes.
    Note      For each cell line, optimal incubation time should be tested experimentally.
  5. Remove the dye working solution and wash cells with 1X Hank’s salt solution and 20 mM Hepes buffer or DPBS once.
  6. Add 100 µL of Adhesion Assay Buffer to the wells.
  7. Monitor the fluorescence intensity using a fluorescence microplate reader at Ex/Em = 490/525 nm (Cutoff = 515 nm). 

Spectrum


Open in Advanced Spectrum Viewer
spectrum

Spectral properties

Excitation (nm)494
Emission (nm)514

References


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The Protein A-mediated binding of Staphylococcus to antibodies in flow cytometric assays and its reduction using FcR blocking reagent.
Authors: Cronin, Ultan P and Girardeaux, Laura and O'Meara, Elaine and Wilkinson, Martin
Journal: Applied and environmental microbiology (2020)
Effect of advanced glycation end product on paraoxonase 2 expression: Its impact on endoplasmic reticulum stress and inflammation in HUVECs.
Authors: Ravi, Ramya and Ragavachetty Nagaraj, Nareshkumar and Subramaniam Rajesh, Bharathidevi
Journal: Life sciences (2020): 117397
Functionalized Graphene Nanoparticles Induce Human Mesenchymal Stem Cells to Express Distinct Extracellular Matrix Proteins Mediating Osteogenesis.
Authors: Newby, Steven D and Masi, Tom and Griffin, Christopher D and King, William J and Chipman, Anna and Stephenson, Stacy and Anderson, David E and Biris, Alexandru S and Bourdo, Shawn E and Dhar, Madhu
Journal: International journal of nanomedicine (2020): 2501-2513
Apolipoprotein M and sphingosine-1-phosphate complex alleviates TNF-α-induced endothelial cell injury and inflammation through PI3K/AKT signaling pathway.
Authors: Liu, Yang and Tie, Li
Journal: BMC cardiovascular disorders (2019): 279
Angiotensin II Type I Receptor Blockade Is Associated with Decreased Cutaneous Scar Formation in a Rat Model.
Authors: Murphy, Amanda and LeVatte, Terry and Boudreau, Colton and Midgen, Craig and Gratzer, Paul and Marshall, Jean and Bezuhly, Michael
Journal: Plastic and reconstructive surgery (2019): 803e-813e
Viable cryopreserved umbilical tissue (vCUT) reduces post-operative adhesions in a rabbit abdominal adhesion model.
Authors: Dhall, Sandeep and Coksaygan, Turhan and Hoffman, Tyler and Moorman, Matthew and Lerch, Anne and Kuang, Jin-Qiang and Sathyamoorthy, Malathi and Danilkovitch, Alla
Journal: Bioactive materials (2019): 97-106
Cardiomyocyte-myofibroblast contact dynamism is modulated by connexin-43.
Authors: Schultz, Francisca and Swiatlowska, Pamela and Alvarez-Laviada, Anita and Sanchez-Alonso, Jose L and Song, Qianqian and de Vries, Antoine A F and Pijnappels, Daniël A and Ongstad, Emily and Braga, Vania M M and Entcheva, Emilia and Gourdie, Robert G and Miragoli, Michele and Gorelik, Julia
Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology (2019): 10453-10468
L-sepiapterin restores SLE serum-induced markers of endothelial function in endothelial cells.
Authors: Jones Buie, Joy N and Pleasant Jenkins, Dorea and Muise-Helmericks, Robin and Oates, Jim C
Journal: Lupus science & medicine (2019): e000294
Allogeneic platelet rich plasma serves as a scaffold for articular cartilage derived chondroprogenitors.
Authors: Vinod, Elizabeth and Vinod Francis, Deepak and Manickam Amirtham, Soosai and Sathishkumar, Solomon and Boopalan, P R J V C
Journal: Tissue & cell (2019): 107-113
Elucidating the expression and function of Numbl during cell adhesion-mediated drug resistance (CAM-DR) in multiple myeloma (MM).
Authors: Huang, Yuejiao and Huang, Xianting and Cheng, Chun and Xu, Xiaohong and Liu, Hong and Yang, Xiaojing and Yao, Li and Ding, Zongmei and Tang, Jie and He, Song and Wang, Yuchan
Journal: BMC cancer (2019): 1269