QCM™ 96-WELL CELL INVASION Assay Kit

Key Applications	Detection Methods
ACT	Fluorescent

Key Applications

Detection Methods

ACT

Fluorescent

Description
Catalogue Number	ECM555
Brand Family	Chemicon®
Trade Name	QCM Chemicon
Description	QCM ECMatrix Cell Invasion Assay, 96-well (8 µm), fluorimetric
Overview	Also available: Cell Comb™ Scratch Assay! Get biochemical data from a scratch assay! Click Here Introduction Invasion through the extracellular matrix (ECM) is an important step in tumor metastasis. Cancer cells initiate invasion by adhering to and spreading along the blood vessel wall. Proteolytic enzymes, such as MMP collagenases, dissolve tiny holes in the sheath-like covering (basement membrane) surrounding the blood vessels to allow cancer cells to invade (Egeblad & Werb, 2002). Microporous membrane inserts are widely used for cell migration and invasion assays. The most widely accepted of which is the Boyden Chamber assay. However, current methods of analysis are time-consuming and tedious, involving cotton swabbing of non-invaded cells on the topside of insert, manual staining and counting. Recently a fluorescence blocking membrane insert was introduced to address these issues; however, this approach requires labeling of the cells with Calcein-AM and extensive washing to remove free Calcein before cell invasion. The effect of this treatment on cell behavior/invasion remains questionable. The CHEMICON QCM™ 96-well Invasion Assay does not require cell labeling, scraping, washing or counting. The 96-well insert and homogenous fluorescence detection format allows for large-scale screening and quantitative comparison of multiple samples. In the CHEMICON QCM™ 96-well Invasion Assay, invaded cells on the bottom of the insert membrane are dissociated from the membrane when incubated with Cell Detachment Buffer. These cells are subsequently lysed and detected by the patented CyQuant GR* dye (Molecular Probes) (Gildea et al., 2000). This green-fluorescent dye exhibits strong fluorescence enhancement when bound to cellular nucleic acids (Jones et al., 2001). *CyQUANT GR® is a registered trademark of Molecular Probes, Inc. The reagent is licensed from Molecular Probes, Inc. and is for use in kits sold by Chemicon International, Inc. for the monitoring of cell invasion and cell migration only. The CHEMICON Cell Invasion Assay Kit provides an efficient system for evaluating the invasion of tumor cells through a basement membrane model. The kit utilizes ECMatrixTM, a reconstituted basement membrane matrix of proteins derived from the Engelbreth Holm-Swarm (EHS) mouse tumor (Albini et al., 1987; Repesh, 1989; Terranova et al., 1986; Liotta, 1984). We examined the kit's performance using human fibrosarcoma (HT-1080) and non-invasive fibroblasts (NIH3T3). The CHEMICON QCM™ 96-well Invasion Assay provides a quick and efficient system for quantitative determination of various factors on cell invasion, including screening of pharmacological agents, evaluation of MMPs or other proteases responsible for cell invasion, or analysis of gene function in transfected cells. In addition, CHEMICON also provides QCM™ 96-well Chemotaxis Cell Migration Assay Systems (8 μm or 3 μm pore), 24-well insert Cell Migration and Invasion Assay Systems, CytoMatrix™ Cell Adhesion strips coated with ECM proteins or anti-integrin antibodies, and QuantiMatrix™ ECM protein ELISA kits. Test Principle: The CHEMICON Cell Invasion Assay is performed in a 96 well invasion plate based on the Boyden chamber principle. This plate contains 96 inserts; each insert contains an 8 μm pore size polycarbonate membrane coated with a thin layer of ECMatrixTM. The ECM layer occludes the membrane pores, blocking non-invasive cells from migrating through. Invasive cells, on the other hand, migrate through the ECM layer and cling to the bottom of the polycarbonate membrane. Invaded cells on the bottom of the insert membrane are dissociated from the membrane when incubated with Cell Detachment Buffer and subsequently lysed and detected by CyQuant GR dye. Application: The CHEMICON Cell Invasion Assay Kit is ideal for evaluation of invasive tumor cells. Each CHEMICON Cell Invasion Assay Kit contains sufficient reagents for the evaluation of 96 samples. The quantitative nature of this assay is especially useful for large scale screening of pharmacological agents. The CHEMICON Cell Invasion Assay Kit is intended for research use only; not for diagnostic or therapeutic applications.
Materials Required but Not Delivered	1. Precision pipettes: sufficient for aliquoting cells. 2. Harvesting buffer: EDTA or trypsin cell detachment buffer. Suggested formulations include a) 2 mM EDTA/PBS, b) 0.05% trypsin in Hanks Balanced Salt Solution (HBSS) containing 25 mM HEPES, or other cell detachment formulations as optimized by individual investigators. Note: Trypsin cell detachment buffer maybe required for difficult cell lines. Allow sufficient time for cell receptor recovery. 3. Tissue culture growth medium appropriate for subject cells, such as DMEM containing 10% FBS. 4. Chemoattractants (eg. 10% FBS) or pharmacological agents for addition to culture medium, if screening is desired. 5. Quenching Medium: serum-free medium, such as DMEM, EMEM, or FBM (fibroblast basal media), containing 5% BSA. Note: Quenching Medium must contain divalent cations (Mg2+, Ca2+) sufficient for quenching EDTA in the harvesting buffer. 6. Sterile PBS or HBSS to wash cells. 7. Distilled water. 8. Low speed centrifuge and tubes for cell harvesting. 9. CO2 incubator appropriate for subject cells. 10. Hemocytometer or other means of counting cells. 11. Trypan blue or equivalent viability stain. 12. Fluorescence plate reader. 13. Sterile cell culture hood.

Description

Catalogue Number

ECM555

Brand Family

Chemicon®

Trade Name

QCM
Chemicon

Description

QCM ECMatrix Cell Invasion Assay, 96-well (8 µm), fluorimetric

Overview

Also available: Cell Comb™ Scratch Assay! Get biochemical data from a scratch assay! Click Here

Introduction
Invasion through the extracellular matrix (ECM) is an important step in tumor metastasis. Cancer cells initiate invasion by adhering to and spreading along the blood vessel wall. Proteolytic enzymes, such as MMP collagenases, dissolve tiny holes in the sheath-like covering (basement membrane) surrounding the blood vessels to allow cancer cells to invade (Egeblad & Werb, 2002).

Microporous membrane inserts are widely used for cell migration and invasion assays. The most widely accepted of which is the Boyden Chamber assay. However, current methods of analysis are time-consuming and tedious, involving cotton swabbing of non-invaded cells on the topside of insert, manual staining and counting. Recently a fluorescence blocking membrane insert was introduced to address these issues; however, this approach requires labeling of the cells with Calcein-AM and extensive washing to remove free Calcein before cell invasion. The effect of this treatment on cell behavior/invasion remains questionable.

The CHEMICON QCM™ 96-well Invasion Assay does not require cell labeling, scraping, washing or counting. The 96-well insert and homogenous fluorescence detection format allows for large-scale screening and quantitative comparison of multiple samples.

In the CHEMICON QCM™ 96-well Invasion Assay, invaded cells on the bottom of the insert membrane are dissociated from the membrane when incubated with Cell Detachment Buffer. These cells are subsequently lysed and detected by the patented CyQuant GR* dye (Molecular Probes) (Gildea et al., 2000). This green-fluorescent dye exhibits strong fluorescence enhancement when bound to cellular nucleic acids (Jones et al., 2001).

*CyQUANT GR® is a registered trademark of Molecular Probes, Inc. The reagent is licensed from Molecular Probes, Inc. and is for use in kits sold by Chemicon International, Inc. for the monitoring of cell invasion and cell migration only.

The CHEMICON Cell Invasion Assay Kit provides an efficient system for evaluating the invasion of tumor cells through a basement membrane model. The kit utilizes ECMatrixTM, a reconstituted basement membrane matrix of proteins derived from the Engelbreth Holm-Swarm (EHS) mouse tumor (Albini et al., 1987; Repesh, 1989; Terranova et al., 1986; Liotta, 1984). We examined the kit's performance using human fibrosarcoma (HT-1080) and non-invasive fibroblasts (NIH3T3).

The CHEMICON QCM™ 96-well Invasion Assay provides a quick and efficient system for quantitative determination of various factors on cell invasion, including screening of pharmacological agents, evaluation of MMPs or other proteases responsible for cell invasion, or analysis of gene function in transfected cells.

In addition, CHEMICON also provides QCM™ 96-well Chemotaxis Cell Migration Assay Systems (8 μm or 3 μm pore), 24-well insert Cell Migration and Invasion Assay Systems, CytoMatrix™ Cell Adhesion strips coated with ECM proteins or anti-integrin antibodies, and QuantiMatrix™ ECM protein ELISA kits.

Test Principle:

The CHEMICON Cell Invasion Assay is performed in a 96 well invasion plate based on the Boyden chamber principle. This plate contains 96 inserts; each insert contains an 8 μm pore size polycarbonate membrane coated with a thin layer of ECMatrixTM. The ECM layer occludes the membrane pores, blocking non-invasive cells from migrating through. Invasive cells, on the other hand, migrate through the ECM layer and cling to the bottom of the polycarbonate membrane. Invaded cells on the bottom of the insert membrane are dissociated from the membrane when incubated with Cell Detachment Buffer and subsequently lysed and detected by CyQuant GR dye.

Application:

The CHEMICON Cell Invasion Assay Kit is ideal for evaluation of invasive tumor cells. Each CHEMICON Cell Invasion Assay Kit contains sufficient reagents for the evaluation of 96 samples. The quantitative nature of this assay is especially useful for large scale screening of pharmacological agents.

The CHEMICON Cell Invasion Assay Kit is intended for research use only; not for diagnostic or therapeutic applications.

Materials Required but Not Delivered

1. Precision pipettes: sufficient for aliquoting cells.

2. Harvesting buffer: EDTA or trypsin cell detachment buffer. Suggested formulations include a) 2 mM EDTA/PBS, b) 0.05% trypsin in Hanks Balanced Salt Solution (HBSS) containing 25 mM HEPES, or other cell detachment formulations as optimized by individual investigators.

Note: Trypsin cell detachment buffer maybe required for difficult cell lines. Allow sufficient time for cell receptor recovery.

3. Tissue culture growth medium appropriate for subject cells, such as DMEM containing 10% FBS.

4. Chemoattractants (eg. 10% FBS) or pharmacological agents for addition to culture medium, if screening is desired.

5. Quenching Medium: serum-free medium, such as DMEM, EMEM, or FBM (fibroblast basal media), containing 5% BSA.

Note: Quenching Medium must contain divalent cations (Mg2+, Ca2+) sufficient for quenching EDTA in the harvesting buffer.

6. Sterile PBS or HBSS to wash cells.

7. Distilled water.

8. Low speed centrifuge and tubes for cell harvesting.

9. CO2 incubator appropriate for subject cells.

10. Hemocytometer or other means of counting cells.

11. Trypan blue or equivalent viability stain.

12. Fluorescence plate reader.

13. Sterile cell culture hood.

Product Information
Components	Sterile 96-well Cell Invasion Plate Assembly: (Part No. 90200) One 96-well feeder tray, and one 96-well Cell Invasion Chamber plate with ECMatrixTM -coated inserts. 96-well Cell Culture Tray: (Part No. 90129) One 96-well feeder tray. Cell Detachment Solution: (Part No. 90131) One bottle - 16 mL. 4X Cell Lysis Buffer: (Part No. 90130) One bottle - 16 mL. CyQuant GR Dye®: (Part No. 90132) One vial - 75 μL
Detection method	Fluorescent

Product Information

Components

Sterile 96-well Cell Invasion Plate Assembly: (Part No. 90200) One 96-well feeder tray, and one 96-well Cell Invasion Chamber plate with ECMatrixTM -coated inserts.
96-well Cell Culture Tray: (Part No. 90129) One 96-well feeder tray.
Cell Detachment Solution: (Part No. 90131) One bottle - 16 mL.
4X Cell Lysis Buffer: (Part No. 90130) One bottle - 16 mL.
CyQuant GR Dye®: (Part No. 90132) One vial - 75 μL

Detection method

Fluorescent

Applications
Application	The CHEMICON Cell Invasion Assay Kit employs a 96-well plate, and provides an efficient system for evaluating the invasion of tumor cells through a basement membrane model.
Key Applications	Activity Assay
Application Notes	Detection of migrated cells is via CyQuant GR Dye®.

Applications

Application

The CHEMICON Cell Invasion Assay Kit employs a 96-well plate, and provides an efficient system for evaluating the invasion of tumor cells through a basement membrane model.

Key Applications

Activity Assay

Application Notes

Detection of migrated cells is via

CyQuant GR Dye®.

Biological Information
Species Reactivity	Human

Biological Information

Species Reactivity

Human

Product Usage Statements
Usage Statement	Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

Product Usage Statements

Usage Statement

Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

Storage and Shipping Information
Storage Conditions	Store kit materials at 2-8°C for up to their expiration date. Do not freeze.

Storage and Shipping Information

Storage Conditions

Store kit materials at 2-8°C for up to their expiration date. Do not freeze.

Packaging Information
Material Size	1 plate
Material Package	96 wells

Packaging Information

Material Size

1 plate

Material Package

96 wells

实验耗材

分子生物学试剂

蛋白与免疫学

生化试剂

细胞及检测试剂

色谱及层析

仪器设备

实验服务

办公用品

QCM™ 96-WELL CELL INVASION Assay Kit

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