Cell Meter™ Cell Adhesion Assay Kit

Effects of Hpgd on the adhesion and angiogenesis of hPAECs. (A) The microscope results showed that OE-Hpgd reduced the adhesion of hPAECs. (B) Statistical histogram of the adherent cells. (C) The microscope results showed that OE-Hpgd reduced the angiogenesis capability. (D) Statistical bar chart of the number of tubules. (**P<0.01). Source: <b>Hpgd affects the progression of hypoxic pulmonary hypertension by regulating vascular remodeling</b> by Meng He, Kelong Tao, Min Xiang & Jian Sun. <em>BMC Pulmonary Medicine</em> volume 23, Article number: 116. April 2023.

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Telephone	1-800-990-8053
Fax	1-800-609-2943
Email	sales@aatbio.com
Quotation	Request
International	See distributors
Shipping	Standard overnight for United States, inquire for international

Spectral properties

Excitation (nm)	494
Emission (nm)	514

Storage, safety and handling

H-phrase	H303, H313, H333
Hazard symbol	XN
Intended use	Research Use Only (RUO)
R-phrase	R20, R21, R22
UNSPSC	12171501

Overview SDS Protocol

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

Excitation	FITC filter set
Emission	FITC filter set
Recommended plate	Black wall/clear bottom

Fluorescence microplate reader

Excitation	490 nm
Emission	525 nm
Cutoff	515 nm
Recommended plate	Black wall/Clear bottom

Components

Example protocol

AT A GLANCE

Protocol summary

Add cells on a plate coated with desired coating material
Incubate cells at 37 °C to allow them to attach
Remove the unattached cells
Add Calcein Ultragreen AM working solution
Incubate the cells at 37 °C for 20-30 minutes
Remove supernatant and wash cells with HHBS or DPBS
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.

Add 100 µL volumes of cells on a plate coated with desired coating material.
Incubate plate at 37 °C for 2 to 3 hours.
Note For each cell line, optimal incubation time should be tested experimentally.
Remove the growth medium and unattached cells.
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.
Remove the dye working solution and wash cells with 1X Hank’s salt solution and 20 mM Hepes buffer or DPBS once.
Add 100 µL of Adhesion Assay Buffer to the wells.
Monitor the fluorescence intensity using a fluorescence microplate reader at Ex/Em = 490/525 nm (Cutoff = 515 nm).

Spectrum

Open in Advanced Spectrum Viewer

Spectral properties

Excitation (nm)	494
Emission (nm)	514

Product Family

Name	Excitation (nm)	Emission (nm)
Cell Meter™ Cell Viability Assay Kit Blue Fluorescence with 405 nm Excitation	406	445
Cell Meter™ Cell Viability Assay Kit Green Fluorescence	494	514
Cell Meter™ Cell Proliferation Assay Kit	503	527

Images

Figure 1. 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.

Figure 2. Effects of Hpgd on the adhesion and angiogenesis of hPAECs. (A) The microscope results showed that OE-Hpgd reduced the adhesion of hPAECs. (B) Statistical histogram of the adherent cells. (C) The microscope results showed that OE-Hpgd reduced the angiogenesis capability. (D) Statistical bar chart of the number of tubules. (**P<0.01). Source: Hpgd affects the progression of hypoxic pulmonary hypertension by regulating vascular remodeling by Meng He, Kelong Tao, Min Xiang & Jian Sun. BMC Pulmonary Medicine volume 23, Article number: 116. April 2023.

Citations

View all 1 citations: Citation Explorer

Hpgd affects the progression of hypoxic pulmonary hypertension by regulating vascular remodeling
Authors: He, Meng and Tao, Kelong and Xiang, Min and Sun, Jian
Journal: BMC Pulmonary Medicine (2023): 1--12

References

View all 50 references: Citation Explorer

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