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id="post-3151" class="post-3151 page type-page status-publish hentry" itemtype="https://schema.org/CreativeWork" itemscope=""> <div class="entry-content entry-content-servises "> <h1>Soft Agar Assay (Colony Formation Assay)</h1> <div class="text-mit-bild-text"> <div class="clearfix text-formatted field field--name-field-text field--type-text-long field--label-hidden field__items"> <p>The Colony Formation Assay, also called a clonogenic assay, allows screening of the therapeutic efficacy of drugs for anchorage-independent cell growth floating in soft agar. In the Soft Agar Assay, cells grow from single cells to cell colonies in an agar solution keeping them from the solid surface and allows growth in an anchorage-independent way.</p> <p>The anchorage-independent formation of cell colonies is one of the hallmark characteristics of cellular transformation and uncontrolled cell growth. Normal epithelial cells are supported by basement membranes providing survival and proliferative signals and undergo apoptosis when placed in suspension culture. Tumor cells, in contrast, evade attachment-regulated apoptosis, leading to uncontrolled proliferation. In order to discover agents that revert cell transformation and inhibit anchorage-independent cell growth, the Soft Agar Assay combines sophisticated 3D cell culture with a high throughput setup and reliable quantification.</p> <h2>What is a Soft Agar Assay?</h2> <p>The Soft Agar Assay is one of two 3D assays offered by <a style="text-decoration: underline!important;" href="/">Reaction Biology</a>. The Soft Agar Assay is used to test the impact of drugs on the proliferation of single cells growing into colonies. In contrast, the <a style="text-decoration: underline!important;" href="/services/cell-based-assays/3d-tumor-spheroid-assay">3D Tumor Spheroid Assay</a> is used for testing a drug’s anti-proliferative and cytotoxic capacity on existing tumor spheroids.</p> <p>Our colony formation assay service provides the following benefits:</p> <ul> <li>Longterm exposure: Up to three weeks of compound exposure possible</li> <li>Available for drug combination treatment</li> <li>All cell lines are subject to authentification via STR profiling</li> </ul> </div> <div class="contact-buttons"> <div class="color-scheme-turquoise block block-block-content block-block-content9594f627-1f8b-4f13-b65e-979c19e399c0"> <a href="https://www.reactionbiology.com/contact/contact-us"> <div class="field field--name-field-icon field--type-image field--label-hidden field__items"> <img decoding="async" src="https://www.reactionbiology.com/wp-content/themes/Divi-child/images/icons-email.svg" alt="E-Mail" title="E-Mail" typeof="foaf:Image"> </div> <div class="field field--name-field-text field--type-string field--label-hidden field__items"> Contact an expert </div> </a> </div> <div class="color-scheme-turquoise block block-block-content block-block-contentc2ce2ae7-66e6-4f1b-87fb-ded5e6bcfb90"> <a class="newsletter-icon-box" href="#"> <div class="field field--name-field-icon field--type-image field--label-hidden field__items"> <img decoding="async" src="https://www.reactionbiology.com/wp-content/themes/Divi-child/images/icons-newsletter.svg" alt="Subscribe to Newsletter" title="Subscribe to Newsletter" typeof="foaf:Image"> </div> <div class="field field--name-field-text field--type-string field--label-hidden field__items"> Subscribe to our newsletter </div> </a> </div> <div class="color-scheme-turquoise block block-block-content block-block-content427638f8-6c52-46af-9064-2fe39ae6579d"> <a href="/#conferences"> <div class="field field--name-field-icon field--type-image field--label-hidden field__items"> <img decoding="async" src="https://www.reactionbiology.com/wp-content/themes/Divi-child/images/icons-meeting.svg" alt="Meet us at your next conference" title="Meet us at your next conference" typeof="foaf:Image"> </div> <div class="field field--name-field-text field--type-string field--label-hidden field__items"> Meet us at your next conference </div> </a> </div> <div class="color-scheme-turquoise block block-block-content block-block-contentff27f1d7-2f17-4dd2-ad72-3ef66b6982de"> <a href="/cdn-cgi/l/email-protection#f6c98583949c939582cbbfd681979882d68299d6859e978493d6829e9f85d6a4939795829f9998d6b49f999a99918fd69a9f989dd6819f829ed68f9983d0979b86cd9499928fcb9e82828685ccd9d9818181d884939795829f9998949f999a99918fd895999bd9859384809f959385d995939a9adb9497859392db978585978f85d9c592db95939a9adb9497859392db978585978f85d985999082db97919784db978585978fdb95999a99988fdb9099849b97829f9998db978585978f"> <div class="field field--name-field-icon field--type-image field--label-hidden field__items"> <img decoding="async" src="https://www.reactionbiology.com/wp-content/themes/Divi-child/images/icons-share.svg" alt="Share" title="Share" typeof="foaf:Image"> </div> <div class="field field--name-field-text field--type-string field--label-hidden field__items"> Share </div> </a> </div> </div> </div> </div> </article> </div> </div> <div id="_anchor" class="paragraph weiss black paragraph--type--paragraph-container paragraph--view-mode--default"><div id="cba-id" class="paragraph text-right images-1 headline-left contact-buttons- paragraph--type--text-with-image paragraph--view-mode--default"><div class="container center"><div class="text-wrapper"><h3>Colony Formation</h3> <div class="text-mit-bild-text"> <div class="clearfix text-formatted field field--name-field-text field--type-text-long field--label-hidden field__items"> <p>Single cells are seeded in soft agar medium for growth independent of the attachment to a surface. Tumor cells, as opposed to other non-transformed cell types, can divide in an anchorage-independent way with single cells building cell colonies over a course of several days or a few weeks.</p> </div> </div> <div class="text-mit-bild-bild field field--name-field-images field--type-image field--label-hidden field__items"> <div class="text-mit-bild-bild-rahmen"> <div class="field__item"> <p style="margin-bottom: 0px;"><img loading="lazy" decoding="async" class="size-full wp-image-1415 aligncenter" src="https://www.reactionbiology.com/wp-content/uploads/2023/07/H460.jpg" alt="H460 cell colonies in the soft agar assay" width="241" height="165" /></p> <div class="caption">H460 cell colonies in the soft agar assay</div> </div> </div> </div> </div></div></div><div id="cba-id2" class="paragraph paragraph--type--tab-container paragraph--view-mode--default"><div class="container center"><h2>Colony Formation Assay Details</h2> <div class="tab-container"><ul class="tab-navigation"><li class="tab-active" data-id="#tab-cba-id221">Assay Description</li><li data-id="#tab-cba-id222">Example Data</li><li data-id="#tab-cba-id223">Screening Details</li></ul><div class="field field--name-field-paragraph field--type-entity-reference-revisions field--label-hidden field__items"><div id="tab-cba-id221" class="tab-content-active paragraph paragraph--type--tab-text-with-image paragraph--view-mode--default"><div class="field field--name-field-title field--type-string field--label-hidden field__items">Assay Description</div><div class="tab-content"><div class="tab-content-wrapper"><div class="clearfix text-formatted field field--name-field-text field--type-text-long field--label-hidden field__items"><p><img decoding="async" class="aligncenter size-full wp-image-1418" src="https://www.reactionbiology.com/wp-content/uploads/2023/07/191022_soft_agar_assay_procedure.png" alt="soft agar assay procedure for colony formation of tumor cells" width="690" /></p> <p>Wells of a 96-well plate were coated with 0.6% soft agar (A), followed by seeding of the cells in 0.4% soft agar (B). After the agar has solidified, compounds are added (B) and cells are incubated several days until colonies have formed (C). A cell viability reagent is added, and fluorescence is measured as an indirect quantification of colony growth in soft agar using Resazurin (D).</p> </div></div></div></div><div id="tab-cba-id222" class=" paragraph paragraph--type--tab-text-with-image paragraph--view-mode--default"><div class="field field--name-field-title field--type-string field--label-hidden field__items">Example Data</div><div class="tab-content"><div class="tab-content-wrapper"><div class="clearfix text-formatted field field--name-field-text field--type-text-long field--label-hidden field__items"><p><img decoding="async" class="aligncenter wp-image-1418" src="https://www.reactionbiology.com/wp-content/uploads/2023/07/Soft-Agar-Example-Data.png" alt="soft agar assay procedure for colony formation of tumor cells" width="579" /></p> <p>The Aurora B inhibitor VX-680 was tested for inhibition of the soft agar growth of A549 non-small cell lung cancer cells at indicated concentrations. Cells were left to form colonies for 5 days, photographed, and stained with a fluorescent cell viability agent. Fluorescence was quantified and, for analysis of IC50 values, was expressed as a percentage of colony growth in the presence of solvent alone.</p> </div></div></div></div><div id="tab-cba-id223" class=" paragraph paragraph--type--tab-text-with-image paragraph--view-mode--default"><div class="field field--name-field-title field--type-string field--label-hidden field__items">Screening Details</div><div class="tab-content"><div class="tab-content-wrapper"><div class="clearfix text-formatted field field--name-field-text field--type-text-long field--label-hidden field__items"><p><strong>Setup:</strong> IC<sub>50</sub> value determination. Drug combination testing is possible.</p> <p><strong>Controls:</strong> DMSO only is high control. Staurosporine treatment (at 1×10-5M) is low control. In every project, we include the IC50 value determination of a reference compound.</p> <p><strong>Cell lines:</strong> Please <a style="text-decoration: underline!imporrtant;" href="/contact/contact-us">inquire</a> to receive a list of cell lines that are established with the Soft Agar Assay format. The establishment of assays with custom cell lines is an option.</p> <p><strong>Report:</strong> A detailed report including assay conditions, methodology, and comprehensive evaluation of data as well as raw data for each analysis will be provided. Photographs of colonies can be provided upon request.</p> <p><strong>Turnaround time</strong> for standard studies is around 3 to 5 weeks. Expedited scheduling and data delivery can be arranged prior to study commencement.</p> <p><strong>Screening facility:</strong> The assay is performed in Freiburg, Germany.</p> <p><strong>Compound requirements:</strong> Compound requirements in brief: 30 µl of the stock solution with 1000x of the highest assay concentration or the equivalent of solid material.</p> <ul class="pdf-listing-1" style="margin-bottom: 15px!important;"> <li class="excel"><a class="custom-asset-link-1" title="FAQ Assay Condition Form" href="https://www.reactionbiology.com/faq/" target="_blank" rel="noopener">Please see our FAQ page to access our cell-based assay condition form (German facility) for further details on compound preparation and shipping.</a></li> </ul> </div></div></div></div></div></div></div></div><div id="cba-id3" class="paragraph paragraph--type--related-resources-container paragraph--view-mode--default"><div class="container center"><h2>Related Resources</h2><div 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