Soft Agar Colony Formation Assay Service

Anchorage-independent cell growth demonstrates the ability of transformed cells to proliferate independently in semi-solid culture medium (e.g. soft agar). It is one of the hallmarks of carcinogenesis. The soft agar colony formation assay is a well-established method used to monitor anchorage-independent growth in vitro, which is considered as the most stringent assay for detecting the tumorigenic potential of transformed cells.

In the soft agar assay, a bottom layer of soft agar is plated and solidified first, followed by seeding of cells in the top soft agar layer. This approach prevents cells from attaching to the bottom of culture plate/dish, allowing transformed cells to form colonies. The principle of this assay suggests that normal adherent cells are unable to grow and divide without anchorage to the extracellular matrix and undergo apoptosis. In contrast, transformed cells are able to grow without adherent condition. Thus, cells capable of forming colonies in soft agar are considered to be transformed and carcinogenic.

Creative Bioarray offers soft agar colony formation assay service to support your research. This assay can be used in cell differentiation, cell transformation, and tumorigenesis studies as well as the efficacy evaluation of anti-tumor therapeutics.

For this assay, cells (treated with test compounds) are cultured with appropriate controls in soft agar medium for 14~28 days. Following this incubation period, newly formed colonies can be analyzed morphologically and quantitatively.

Figure 1. Schematic Overview of Soft Agar Colony Formation Assay Protocol.

Assay Procedure

• Coat the plate with the bottom agar layer
• Plate the top agar layer containing cells, followed by drug treatment
• Incubate for 14-28 days and feed cells twice a week
• Count and analyze the colonies larger than 70 µm

Figure 2. Images (A) and Colony Count (B) for MCF10DCIS Cells with or without BB-CLA Treatment in Soft Agar.

Advantages

• Fast, convenient, and accurate
• Cell lines and conditions can be established upon request
• Finding transformation targets
• Allowing profiling of test compounds in physiological relevant environment

Study Example

Figure 3. Images for the test cell line and positive and negative control cell lines with and without the addition of bortezomib at 1 µM in the soft agar assay (scale bar, 70 µm).

Contact Us Today

If you are interested in our soft agar colony formation assay and would like to find out more, please contact us today. Our scientists would be happy to answer your questions. We look forward to hearing from you. Reach us now for support.

References

1. Borowicz S, et al. The soft agar colony formation assay. J Vis Exp. 2014;(92):e51998. Published 2014 Oct 27. doi:10.3791/51998.
2. Horibata S, et al. Utilization of the Soft Agar Colony Formation Assay to Identify Inhibitors of Tumorigenicity in Breast Cancer Cells. J Vis Exp. 2015;(99):e52727. Published 2015 May 20. doi:10.3791/52727.
3. Anderson SN, et al. A high-throughput soft agar assay for identification of anticancer compound. J Biomol Screen. 2007 Oct;12(7):938-45. doi: 10.1177/1087057107306130.

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For research use only. Not for any other purpose.