Flow CAST- Assay Overview Video

Video Transcript:

Hello, I’m Michele Romano, product manager at BUHLMANN for the CAST^® product line.

Today, let me show you the Flow CAST^® kit and the CAST^® allergens, our products for Basophil Activation Test, the allergy test in a vial.

BAT is a combination of specifically stimulated basophils from peripheral blood and the detection of activation with flow cytometry.  BÜHLMANN has been at the forefront of Basophil Activation Test development for over 20 years, combining innovation with a commitment to high-quality standards.

Flow CAST^® is the result of an extensive process of development, validation, and simplification of the assay. Our focus is to offer a product that is robust and user-friendly at the same time.

Opening a kit, you can find the first key reagent is our stimulation buffer. We have optimized it to work with basophils in EDTA whole blood and we added IL3 for the optimal activation of CD63.  We are using CD63 as the activation marker and CCR3 as the selection marker of basophils in our gating strategy to identify activated basophils.  Also, antibodies anti CD63, FITC conjugated, and anti CCR3-PE are our pre-mixed, ready to use solution in the staining reagent vial.

We added two stimulation controls: a monoclonal Ab for the alpha subunit of the high-affinity receptor for IgE and the fMLP, for the  perfect combination of positive controls to discriminate between the described IgE non-responder from samples which could just have some problems in making the assay.

(Additional information: Approximately 5% of individuals tested have peripheral blood basophils that do not respond to IgE receptor stimulation- these are considered IgE non-responders.  So it is not possible to evaluate an allergen IgE mediated activation with BAT testing in these individuals.  Therefore the addition of a positive activation induced by a second independent activator (fMLP) with its own specific basophil receptor is necessary to discriminate real IgE-non responders from a specific non-activation status of basophils(e.g. lab handling or pre-analytical aspects).  In other words, by adding fMLP, the basophils in IgE non-responders are activated in vitro, demonstrating that the stimulation is specific to the receptor (real IgE non responders) rather than a result of the specimen handling.)

We are proud to be the unique company offering a huge list of high-quality allergens specifically developed for the use in BAT, comprising several fields of allergy: drug, food, inhalants, and hymenoptera allergies. To note within food allergens, we can offer a full panel of allergens relevant for pediatric food allergies, like milk egg, peanut, and nuts.

Our experience in BAT testing suggests that CD63 is the preferred activation marker. We are also offering a separate reagent set for CD203c which includes a staining reagent with anti-CD203c Ab and a stimulation buffer that does not contain IL3.

For technical resources, more information on products and our allergens you can refer to our websites buhlmannlabs.com and buhlamannlabs.ch, where you can also find some news, updates and upcoming events relevant to our Basophil Activation Test.