Drosophila Spike-In Control Kit for CUT&Tag (Rabbit) #29811
Product Information
Storage
Product Description
The Drosophila Spike-In Control Kit for CUT&Tag (Rabbit) is conveniently designed to provide the reagents required to allow for normalization of variabilities across the entire CUT&Tag experiment using rabbit primary antibodies. It starts with the addition of cryopreserved Drosophila spike-in nuclei to cell samples prior to performing the CUT&Tag assay. Like the cells of interest, the spike-in nuclei also bind to the Concanavalin A beads, providing a method to immobilize the spike-in nuclei for subsequent washes and incubations. During the antibody incubation step, an H2Av Rabbit Monoclonal Antibody is added to the reaction in addition to the rabbit primary antibody against the target of interest. This H2Av Rabbit Monoclonal Antibody provides a reliable method to target, digest, and enrich histone H2Av-bound Drosophila chromatin in a consistent way across all samples. This enrichment of H2Av-bound Drosophila chromatin occurs in the same tube as the enrichment of the target of interest from the test cell chromatin, ensuring that the spike-in control nuclei are subjected to all of the same steps and conditions as the test cells. A normalization factor is generated based on the Drosophila chromatin enrichment signal and is applied to the test genome to normalize signals across samples (see Figures 1-4). Thus, this spike-in normalization approach enables reliable and consistent normalization across samples, correcting for variations in starting cell number, technical variation among samples during processing, and nonspecific enrichment coming from isotype control antibodies (i.e., nonspecific rabbit IgG, see Figure 5).
This kit is intended for use only in combination with our CUT&Tag Assay Kit #77552 or other validated in-house CUT&Tag protocols. It provides three 165 µL vials of cryopreserved Drosophila Spike-In Nuclei Control. Each vial contains sufficient nuclei for eight CUT&Tag reactions using 100,000-250,000 cells or 1–2.5 mg of tissue, when used at the recommended volume. Also included is the H2Av Rabbit Monoclonal Antibody, which specifically detects the Drosophila histone variant H2Av, showing no cross-reactivity to mammalian histones.
If CUT&Tag assays are performed with mouse primary antibodies, DO NOT use this kit. Please refer to our Drosophila Spike-In Control Kit for CUT&Tag (Mouse) #19629 for sample normalizations.
Specificity / Sensitivity
Background
A major challenge in comparing CUT&Tag datasets is being able to account for differences in signal resulting from sample-to-sample variability, such as differences in starting cell number, technical variations among samples during processing, and variations in global histone modifications. The Drosophila Spike-In Control Kit for CUT&Tag (Rabbit) provides a complete normalization strategy that allows for normalization of variabilities across the entire experiment (4,5), ensuring a more accurate measurement of protein–DNA interactions between samples and across experiments.
Limited Uses
Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.
Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.