

SNAP-CUTANA™ K-MetStat Panel
{"url":"https://www.epicypher.com/products/nucleosomes/snap-cutana-k-metstat-panel","add_this":[{"service":"facebook","annotation":""},{"service":"email","annotation":""},{"service":"print","annotation":""},{"service":"twitter","annotation":""},{"service":"linkedin","annotation":""}],"gtin":null,"id":"883","bulk_discount_rates":[],"can_purchase":true,"meta_description":"16 barcoded spike-in nucleosomes provide quantitative sample normalization and confirmation of K-methyl antibody specificity in CUT&RUN and CUT&Tag","category":["Nucleosomes","Nucleosomes/SNAP-CUTANA™ Spike-in Controls","Epigenetics Kits and Reagents/CUTANA™ ChIC / CUT&RUN Assays","Epigenetics Kits and Reagents/CUTANA™ CUT&Tag Assays"],"AddThisServiceButtonMeta":"","main_image":{"data":"https://cdn11.bigcommerce.com/s-y9o92/images/stencil/{:size}/products/883/911/CUTANAspike-inthumbnail_RGB_white_w_border__29403.1634679576.png?c=2","alt":"SNAP-CUTANA™ K-MetStat Panel"},"add_to_wishlist_url":"/wishlist.php?action=add&product_id=883","shipping":{"calculated":true},"num_reviews":0,"weight":"0.00 LBS","custom_fields":[{"id":"1008","name":"Pack Size","value":"50 Reactions"},{"id":"1009","name":"Internal Comment","value":"Excess in bottom shelf of Venom."}],"sku":"19-1002","description":"<div class=\"service_accordion product-droppdown\">\n <div class=\"container\">\n <div id=\"prodAccordion\">\n <div id=\"ProductDescription\" class=\"Block Panel current\">\n <h3 class=\"sub-title1\">Description</h3>\n <div\n class=\"ProductDescriptionContainer product-droppdown__section-description-specific\"\n >\n <p>\n The SNAP-CUTANA K-MetStat Panel of spike-in controls for CUT&RUN and\n CUT&Tag offers an all-in-one solution to determine antibody\n specificity for histone posttranslational modifications (PTMs),\n monitor assay success, and normalize data for quantitative chromatin\n mapping. The panel contains designer nucleosomes (dNucs)\n representing 16 different K-methyl PTM states: mono-, di-, and\n trimethylation at H3K4, H3K9, H3K27, H3K36, & H4K20, as well as\n unmodified control (<strong>Figure 1</strong>). Each PTM is\n represented by two unique DNA-barcoded templates (A and B, for an\n internal technical replicate). Each dNuc is individually conjugated\n to paramagnetic beads and pooled into a single panel for convenient\n one-step spike-in to CUT&RUN and CUT&Tag experiments. The panel is\n added to samples alongside ConA-immobilized cells prior to the\n addition of an antibody targeting a histone lysine methylation state\n or IgG negative control (see <strong>Application Notes</strong> and\n <strong>Table 1</strong>). pAG-MNase-mediated release or\n pAG-Tn5-mediated tagmentation of genomic chromatin and the barcoded\n nucleosomes is dependent on the specificity of the antibody used.\n After sequencing, the relative read count of each spike-in\n nucleosome barcode provides a quantitative metric of on- vs.\n off-target recovery (<strong>Figures 2 and 3</strong>) as well as\n quantitative sample normalization, thereby gauging experimental\n success, guiding troubleshooting efforts, and enabling reliable\n cross-sample comparisons.\n </p>\n <p\n style=\"\n background-color: #4698cb;\n color: #fff;\n padding: 1.3rem;\n text-align: center;\n border-radius: 12px;\n \"\n >\n <a\n target=\"_blank\"\n style=\"color: #fff; text-decoration: none\"\n href=\"/products/nucleosomes/snap-cutana-k-metstat-panel#userguide\"\n >The SNAP-CUTANA Spike-in User Guide - everything you need to know\n in one place\n </a>\n </p>\n </div>\n </div>\n </div>\n <div id=\"prodAccordion\">\n <div id=\"ProductDescription\" class=\"Block Panel current\">\n <h3 class=\"sub-title1\">Validation Data</h3>\n <div\n class=\"ProductDescriptionContainer product-droppdown__section-description-specific\"\n >\n <section class=\"image-picker\">\n <div class=\"image-picker__left\">\n <div\n class=\"image-picker__main-content_active image-picker__main-content\"\n >\n <div class=\"image-picker__header-content\">\n <button class=\"image-picker__left-arrow\">\n <svg\n class=\"image-picker__svg-left\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M16.67 0l2.83 2.829-9.339 9.175 9.339 9.167-2.83 2.829-12.17-11.996z\"\n />\n </svg>\n </button>\n <a\n href=\"/content/images/products/nucleosomes/19-1002_graphic.jpeg\"\n target=\"_blank\"\n class=\"image-picker__main-image-link\"\n ><img loading=\"lazy\"\n alt=\"19-1002_graphic\"\n src=\"/content/images/products/nucleosomes/19-1002_graphic.jpeg\"\n class=\"image-picker__main-image\"\n />\n <span class=\"image-picker__main-image-caption\"\n >(Click to enlarge)</span\n ></a\n >\n <button class=\"image-picker__right-arrow\">\n <svg\n class=\"image-picker__svg-right\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M7.33 24l-2.83-2.829 9.339-9.175-9.339-9.167 2.83-2.829 12.17 11.996z\"\n />\n </svg>\n </button>\n </div>\n <p>\n <span class=\"image-picker__span-content\"\n ><strong\n >Figure 1: Schematic of SNAP-CUTANA Spike-in\n Controls</strong\n ><br />\n The K-MetStat Panel controls for 15 widely studied methyl\n states (see Description) and an unmodified control (me0),\n with each of the 16 octamers wrapped with two uniquely\n barcoded DNA templates (A and B). Each 250 bp DNA template\n contains a 123 bp 601 nucleosome positioning sequence (gray)\n [1], a unique 22 bp DNA-barcode (white; 32 barcodes total),\n and a 5’ biotin-TEG. The 5’ and 3’ linkers (blue) are\n compatible with cleavage by pAG-MNase (EpiCypher\n <a\n href=\"https://www.epicypher.com/products/epigenetics-reagents-and-assays/cutana-chic-cut-run-kit\"\n >14-1048</a\n >,\n <a\n href=\"https://www.epicypher.com/products/epigenetics-reagents-and-assays/cutana-pag-mnase-for-chic-cut-and-run-workflows-50-rxns\"\n >15-1016</a\n >) during CUT&RUN as well as tagmentation by pAG-Tn5\n (EpiCypher\n <a\n href=\"https://www.epicypher.com/products/featured-products/cutana-pag-tn5-for-cut-tag-50-rxns\"\n >15-1017</a\n >\n ) during CUT&Tag. The dNucs are individually pre-conjugated\n to paramagnetic beads and pooled for convenient use.\n </span>\n </p>\n </div>\n <div class=\"image-picker__main-content\">\n <div class=\"image-picker__header-content\">\n <button class=\"image-picker__left-arrow\">\n <svg\n class=\"image-picker__svg-left\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M16.67 0l2.83 2.829-9.339 9.175 9.339 9.167-2.83 2.829-12.17-11.996z\"\n />\n </svg>\n </button>\n <a\n href=\"/content/images/products/nucleosomes/KmetStat_Fig2_CUTRUN_Heat.jpeg\"\n target=\"_blank\"\n class=\"image-picker__main-image-link\"\n ><img loading=\"lazy\"\n alt=\"KmetStat_Fig2_CUTRUN_Heat\"\n src=\"/content/images/products/nucleosomes/KmetStat_Fig2_CUTRUN_Heat.jpeg\"\n class=\"image-picker__main-image\"\n />\n <span class=\"image-picker__main-image-caption\"\n >(Click to enlarge)</span\n ></a\n >\n <button class=\"image-picker__right-arrow\">\n <svg\n class=\"image-picker__svg-right\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M7.33 24l-2.83-2.829 9.339-9.175-9.339-9.167 2.83-2.829 12.17 11.996z\"\n />\n </svg>\n </button>\n </div>\n <p>\n <span class=\"image-picker__span-content\"\n ><strong\n >Figure 2: Panel Identity Profiling by CUT&RUN </strong\n ><br />\n CUT&RUN was performed on 500k fresh K-562 cells utilizing\n best in class identified antibodies to each PTM represented\n in the SNAP-CUTANA K-MetStat Panel. Prior to antibody\n addition, 2 µL of the control panel was spiked into each\n sample. CUT&RUN sequencing reads were aligned to the unique\n DNA barcodes in the panel and normalized to either on-target\n (anti-K-methyl PTM) or total counts (IgG, Input). Antibody\n enrichment of the expected spike-in nucleosome target (red)\n confirmed the identity and integrity of each member of the\n panel. CUT&RUN with IgG control antibody showed no\n preferential enrichment for any particular nucleosome\n (blue), as expected. Direct sequencing of the panel (Input)\n indicated balanced pooling of each nucleosome. While\n EpiCypher has not yet identified an H4K20me1 antibody with a\n preferred specificity profile in CUT&RUN, the clone shown\n has sufficient PTM preference to confirm the integrity of\n the barcoded nucleosome.\n <br />\n <br />\n <strong>Note:</strong> While most antibodies used in these\n experiments show good target specificity in our\n <strong>standard</strong> CUT&RUN / CUT&Tag conditions,\n changes to antibody lots and experimental variables (cell\n number, cell type, experimental treatment, salt\n concentration, etc.) can have adverse effects on antibody\n specificity.\n <strong\n >Single-point antibody validation under optimal conditions\n is not a suitable substitute for controlled experiments\n with spike-ins.</strong\n >\n EpiCypher recommends using SNAP spike-in controls to monitor\n experimental success and accuracy in every possible\n reaction.\n </span>\n </p>\n </div>\n <div class=\"image-picker__main-content\">\n <div class=\"image-picker__header-content\">\n <button class=\"image-picker__left-arrow\">\n <svg\n class=\"image-picker__svg-left\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M16.67 0l2.83 2.829-9.339 9.175 9.339 9.167-2.83 2.829-12.17-11.996z\"\n />\n </svg>\n </button>\n <a\n href=\"/content/images/products/nucleosomes/19-1002_CUTTag_heat.jpeg\"\n target=\"_blank\"\n class=\"image-picker__main-image-link\"\n >\n <img loading=\"lazy\"\n alt=\"19-1002_CUTTag_heat\"\n src=\"/content/images/products/nucleosomes/19-1002_CUTTag_heat.jpeg\"\n class=\"image-picker__main-image\"\n />\n <span class=\"image-picker__main-image-caption\"\n >(Click to enlarge)</span\n >\n </a>\n <button class=\"image-picker__right-arrow\">\n <svg\n class=\"image-picker__svg-right\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M7.33 24l-2.83-2.829 9.339-9.175-9.339-9.167 2.83-2.829 12.17 11.996z\"\n />\n </svg>\n </button>\n </div>\n <p>\n <span class=\"image-picker__span-content\">\n <strong\n >Figure 3: Panel Identity Profiling by CUT&Tag</strong\n ><br />\n CUT&Tag was performed on 100k fresh K-562 cells utilizing\n best in class identified antibodies to each PTM represented\n in the SNAP-CUTANA K-MetStat Panel. Prior to antibody\n addition, 2 µL of the control panel was spiked into each\n sample. CUT&Tag sequencing reads were aligned to the unique\n DNA barcodes in the panel and normalized to either on-target\n (anti-K-methyl PTM) or total counts (IgG). Antibody\n enrichment of the expected spike-in nucleosome target (red)\n confirmed the identity and integrity of each member of the\n panel. CUT&Tag with IgG control antibody showed no\n preferential enrichment for any particular nucleosome\n (blue), as expected.\n <br />\n <br />\n <strong>Note:</strong> While most antibodies used in these\n experiments show good target specificity in our\n <strong>standard</strong> CUT&RUN / CUT&Tag conditions,\n changes to antibody lots and experimental variables (cell\n number, cell type, experimental treatment, salt\n concentration, etc.) can have adverse effects on antibody\n specificity.\n <strong\n >Single-point antibody validation under optimal conditions\n is not a suitable substitute for controlled experiments\n with spike-ins.</strong\n >\n EpiCypher recommends using SNAP spike-in controls to monitor\n experimental success and accuracy in every possible\n reaction.\n </span>\n </p>\n </div>\n <div class=\"image-picker__main-content\">\n <div class=\"image-picker__header-content\">\n <button class=\"image-picker__left-arrow\">\n <svg\n class=\"image-picker__svg-left\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M16.67 0l2.83 2.829-9.339 9.175 9.339 9.167-2.83 2.829-12.17-11.996z\"\n />\n </svg>\n </button>\n <a\n href=\"/content/images/products/nucleosomes/19-1002_Panel_Dilution.jpeg\"\n target=\"_blank\"\n class=\"image-picker__main-image-link\"\n >\n <img loading=\"lazy\"\n alt=\"19-1002_Panel_Dilution\"\n src=\"/content/images/products/nucleosomes/19-1002_Panel_Dilution.jpeg\"\n class=\"image-picker__main-image\"\n />\n <span class=\"image-picker__main-image-caption\"\n >(Click to enlarge)</span\n >\n </a>\n <button class=\"image-picker__right-arrow\">\n <svg\n class=\"image-picker__svg-right\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M7.33 24l-2.83-2.829 9.339-9.175-9.339-9.167 2.83-2.829 12.17 11.996z\"\n />\n </svg>\n </button>\n </div>\n <p>\n <span class=\"image-picker__span-content\">\n <strong\n >Table 1: Recommended SNAP-CUTANA Spike-In Dilution for\n CUT&RUN / CUT&Tag Samples of Varying Starting Cell\n Number</strong\n ><br />\n </span>\n </p>\n </div>\n <div class=\"image-picker__main-content\">\n <div class=\"image-picker__header-content\">\n <button class=\"image-picker__left-arrow\">\n <svg\n class=\"image-picker__svg-left\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M16.67 0l2.83 2.829-9.339 9.175 9.339 9.167-2.83 2.829-12.17-11.996z\"\n />\n </svg>\n </button>\n <a\n href=\"/content/images/products/nucleosomes/19-1002_Native_TDS.jpeg\"\n target=\"_blank\"\n class=\"image-picker__main-image-link\"\n >\n <img loading=\"lazy\"\n alt=\"19-1002_Native_TDS\"\n src=\"/content/images/products/nucleosomes/19-1002_Native_TDS.jpeg\"\n class=\"image-picker__main-image\"\n />\n <span class=\"image-picker__main-image-caption\"\n >(Click to enlarge)</span\n >\n </a>\n <button class=\"image-picker__right-arrow\">\n <svg\n class=\"image-picker__svg-right\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M7.33 24l-2.83-2.829 9.339-9.175-9.339-9.167 2.83-2.829 12.17 11.996z\"\n />\n </svg>\n </button>\n </div>\n <p>\n <span class=\"image-picker__span-content\">\n <strong>Figure 4: DNA Gel Data</strong><br />\n Representative images for SNAP-CUTANA K-MetStat nucleosomes\n resolved by native PAGE and stained with ethidium bromide to\n confirm intact nucleosome assembly with minimal free DNA.\n Lane 1: Free 250 bp DNA used in nucleosome assembly (100\n ng). Lane 2: Intact nucleosomes (400 ng). Comparable\n experiments were performed for the entire SNAP-CUTANA\n K-MetStat Panel.\n </span>\n </p>\n </div>\n <div class=\"image-picker__main-content\">\n <div class=\"image-picker__header-content\">\n <button class=\"image-picker__left-arrow\">\n <svg\n class=\"image-picker__svg-left\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M16.67 0l2.83 2.829-9.339 9.175 9.339 9.167-2.83 2.829-12.17-11.996z\"\n />\n </svg>\n </button>\n <a\n href=\"/content/images/products/nucleosomes/19-1002_Octamer_TDS.jpeg\"\n target=\"_blank\"\n class=\"image-picker__main-image-link\"\n >\n <img loading=\"lazy\"\n alt=\"19-1002_Octamer_TDS\"\n src=\"/content/images/products/nucleosomes/19-1002_Octamer_TDS.jpeg\"\n class=\"image-picker__main-image\"\n />\n <span class=\"image-picker__main-image-caption\"\n >(Click to enlarge)</span\n >\n </a>\n <button class=\"image-picker__right-arrow\">\n <svg\n class=\"image-picker__svg-right\"\n width=\"24\"\n height=\"24\"\n viewBox=\"0 0 24 24\"\n >\n <path\n d=\"M7.33 24l-2.83-2.829 9.339-9.175-9.339-9.167 2.83-2.829 12.17 11.996z\"\n />\n </svg>\n </button>\n </div>\n <p>\n <span class=\"image-picker__span-content\">\n <strong>Figure 5: Protein Gel Data</strong><br />\n Representative Coomassie stained PAGE of SNAP-CUTANA\n K-MetStat dNucs (1 µg each) demonstrates the purity of\n histones in the preparation. Sizes of molecular weight\n markers and positions of the core histones (H2A, H2B, H3 and\n H4) are indicated. Comparable experiments were performed for\n the entire SNAP-CUTANA K-MetStat Panel.\n </span>\n </p>\n </div>\n </div>\n <aside class=\"image-picker__right\">\n <div class=\"image-picker__gallery\">\n <img loading=\"lazy\"\n alt=\"19-1002_graphic\"\n src=\"/content/images/products/nucleosomes/19-1002_graphic.jpeg\"\n width=\"200\"\n class=\"image-picker__side-image image-picker__side-image_active\"\n role=\"button\"\n />\n <img loading=\"lazy\"\n alt=\"KmetStat_Fig2_CUTRUN_Heat\"\n src=\"/content/images/products/nucleosomes/KmetStat_Fig2_CUTRUN_Heat.jpeg\"\n class=\"image-picker__side-image\"\n role=\"button\"\n />\n <img loading=\"lazy\"\n alt=\"19-1002_CUTTag_heat\"\n src=\"/content/images/products/nucleosomes/19-1002_CUTTag_heat.jpeg\"\n class=\"image-picker__side-image\"\n role=\"button\"\n />\n <img loading=\"lazy\"\n alt=\"19-1002_Panel_Dilution\"\n src=\"/content/images/products/nucleosomes/19-1002_Panel_Dilution.jpeg\"\n class=\"image-picker__side-image\"\n role=\"button\"\n />\n <img loading=\"lazy\"\n alt=\"19-1002_Native_TDS\"\n src=\"/content/images/products/nucleosomes/19-1002_Native_TDS.jpeg\"\n class=\"image-picker__side-image\"\n role=\"button\"\n />\n <img loading=\"lazy\"\n alt=\"19-1002_Octamer_TDS\"\n src=\"/content/images/products/nucleosomes/19-1002_Octamer_TDS.jpeg\"\n class=\"image-picker__side-image\"\n role=\"button\"\n />\n </div>\n </aside>\n </section>\n </div>\n </div>\n </div>\n <div id=\"userguide\"></div>\n <div id=\"prodAccordion\">\n <div id=\"ProductDescription\" class=\"Block Panel\">\n <h3 class=\"sub-title1\">Technical Information</h3>\n <div\n class=\"ProductDescriptionContainer product-droppdown__section-description\"\n >\n <div class=\"product-tech-info\">\n <div class=\"product-tech-info__line-item\">\n <div class=\"product-tech-info__line-item-left\">\n <b>Formulation</b>\n </div>\n <div class=\"product-tech-info__line-item-right\">\n A mixture of 16 PTM-defined semi-synthetic nucleosomes\n conjugated to paramagnetic beads in 10 mM sodium cacodylate pH\n 7.5, 100 mM NaCl, 1 mM EDTA, 50% glycerol (w/v), 1x Protease\n Inhibitor Cocktail, 100 μg/mL BSA, 10 mM β-mercaptoethanol.\n Molarity = 3 nM each, 96 nM total nucleosome. Average MW =\n ~263,072.7 Da.\n </div>\n </div>\n <div class=\"product-tech-info__line-item\">\n <div class=\"product-tech-info__line-item-left\">\n <b>Storage and Stability</b>\n </div>\n <div class=\"product-tech-info__line-item-right\">\n Store at -20°C.\n <strong\n >Lower temperatures can cause freezing and will permanently\n damage the magnetic beads</strong\n >. Stable for six months from date of receipt.\n </div>\n </div>\n </div>\n </div>\n </div>\n </div>\n <div id=\"prodAccordion\">\n <div id=\"ProductDescription\" class=\"Block Panel\">\n <h3 class=\"sub-title1\">Application Notes</h3>\n <div\n class=\"ProductDescriptionContainer product-droppdown__section-description\"\n >\n <p>\n <em\n >See the most recent\n <a href=\"/resources/protocols\"\n >EpiCypher CUT&RUN or CUT&Tag protocols</a\n >\n for detailed information on workflow integration, expected\n results, data analysis, and troubleshooting. In Brief:</em\n >\n </p>\n <p>\n <u><strong>Product Use:</strong></u> Use the SNAP-CUTANA K-MetStat\n Panel for control reactions containing positive (e.g.\n <a\n href=\"/products/antibodies/snap-chip-certified-antibodies/histone-h3k4me3-antibody-snap-chip-certified-cutana-cut-run-compatible\"\n >H3K4me3</a\n >) and negative (<a\n href=\"/products/epigenetics-reagents-and-assays/cutana-chic-cut-run-assays/cutana-rabbit-igg-cut-run-negative-control-antibody\"\n >IgG</a\n >) antibodies, as well as samples with an antibody to any of the 15\n lysine methyl states in the K-MetStat Panel. Just before antibody\n addition, spike in 2 μL per 500k cells for CUT&RUN and 2 μL per 100k\n cells for CUT&Tag. If using less than the standard number of cells,\n decrease the amount of SNAP-CUTANA spike-in linearly by preparing a\n “working stock” dilution of the panel in the appropriate buffer,\n made fresh the day of use (<strong>Table 1</strong>). Adjust\n spike-in volume as needed; aiming for the spike-ins to comprise ~1%\n of the total sequencing reads. Expect higher for low abundance\n targets / negative controls (IgG; ~10-20%) and lower for high\n abundance targets (H3K27me3; 0.1-1%). <strong>Table 1</strong> gives\n recommended dilution amounts for varying numbers of starting cells,\n but optimization may be required for user-specific conditions.\n </p>\n\n <p>\n <u><strong>Data Analysis:</strong></u> Perform paired end sequencing\n for a minimum of 50 bases. The Widom 601 DNA and DNA barcodes are\n distinct from human, mouse, fly, and yeast genomes such that they\n can be readily distinguished from sample chromatin. A shell script\n (.sh file extension) for spike-in alignment and an excel template\n for heatmap generation are available in the\n <strong>Documents & Resources</strong>\n section below. The shell script can be opened with any basic text\n editor program and contains detailed instructions hashed (#) at the\n beginning of the document. Make sure to copy and paste the R1 & R2\n echo loop so there is a set for each sample being analyzed.\n </p>\n </div>\n </div>\n </div>\n <div id=\"prodAccordion\">\n <div id=\"ProductDescription\" class=\"Block Panel\">\n <h3 class=\"sub-title1\">References</h3>\n <div\n class=\"ProductDescriptionContainer product-droppdown__section-description\"\n >\n <strong>Background References:</strong>\n <br />\n [1] Lowary & Widom <em>J. Mol. Biol.</em> (1998). PMID:\n <a\n href=\"https://pubmed.ncbi.nlm.nih.gov/9514715/\"\n target=\"_blank\"\n title=\"New DNA sequence rules for high affinity binding to histone octamer and sequence-directed nucleosome positioning\"\n >9514715</a\n ><br />\n </div>\n </div>\n </div>\n <div id=\"prodAccordion\">\n <div id=\"ProductDescription\" class=\"Block Panel current\">\n <h3 class=\"sub-title1\">Documents & Resources</h3>\n\n <div\n class=\"ProductDescriptionContainer product-droppdown__section-description-specific\"\n >\n <div class=\"product-documents\">\n <a\n href=\"/content/documents/tds/19-1002.pdf\"\n target=\"_blank\"\n class=\"product-documents__link\"\n >\n <svg\n version=\"1.1\"\n id=\"Layer_1\"\n xmlns=\"http://www.w3.org/2000/svg\"\n xmlns:xlink=\"http://www.w3.org/1999/xlink\"\n x=\"0px\"\n y=\"0px\"\n viewBox=\"0 0 228 240\"\n style=\"enable-background: new 0 0 228 240\"\n xml:space=\"preserve\"\n class=\"product-documents__icon\"\n alt=\"16-0030 Datasheet\"\n >\n <g>\n <path\n 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c2.02,3.63,3.04,7.98,3.04,13.04c0,5.06-1,9.42-3,13.08c-2,3.66-4.91,6.45-8.73,8.38c-3.82,1.93-8.4,2.9-13.73,2.9H69.92V125.68z\n M88.07,165.63c10.35,0,15.52-5.22,15.52-15.66c0-10.4-5.17-15.59-15.52-15.59h-7.38v31.26H88.07z\"\n />\n <path\n class=\"product-documents__svg-pdf\"\n d=\"M122.57,125.68h32.84v8.49h-22.22v11.18h20.84v8.49h-20.84v20.49h-10.63V125.68z\"\n />\n </g>\n </svg>\n </a>\n <a\n class=\"product-documents__link\"\n href=\"/content/documents/SNAP-CUTANA_K-MetStat_user_guide.pdf\"\n ><span class=\"product-documents__info\" s\n >SNAP-CUTANA™ Spike-in User Guide for CUT&RUN and\n CUT&Tag</span\n >\n </a>\n </div>\n </div>\n <div class=\"product-documents\">\n <a\n href=\"/content/documents/SNAP-CUTANA_K-MetStat_Panel_ShellScript.sh\"\n target=\"_blank\"\n class=\"product-documents__link\"\n >\n <svg\n version=\"1.1\"\n id=\"Layer_1\"\n xmlns=\"http://www.w3.org/2000/svg\"\n xmlns:xlink=\"http://www.w3.org/1999/xlink\"\n x=\"0px\"\n y=\"0px\"\n viewBox=\"0 0 228 240\"\n style=\"enable-background: new 0 0 228 240\"\n xml:space=\"preserve\"\n class=\"product-documents__icon\"\n >\n <g>\n <path\n class=\"product-documents__svg-txt\"\n d=\"M199.12,66.39l-51.16-51.16c-1.43-1.43-3.35-2.23-5.37-2.23H41.61c-4.2,0-7.61,3.41-7.61,7.61v197.79\n c0,4.2,3.41,7.61,7.61,7.61h152.14c4.2,0,7.61-3.41,7.61-7.61V71.79C201.36,69.77,200.55,67.82,199.12,66.39L199.12,66.39z\n M183.81,75.28h-44.74V30.54L183.81,75.28z M184.24,208.88H51.12V30.12h71.79v51.35c0,5.51,4.47,9.98,9.98,9.98h51.35V208.88z\n M115.78,144.7H72.04c-1.05,0-1.9,0.86-1.9,1.9v11.41c0,1.05,0.86,1.9,1.9,1.9h43.74c1.05,0,1.9-0.86,1.9-1.9V146.6\n C117.68,145.55,116.82,144.7,115.78,144.7z M70.13,114.27v11.41c0,1.05,0.86,1.9,1.9,1.9h91.29c1.05,0,1.9-0.86,1.9-1.9v-11.41\n c0-1.05-0.86-1.9-1.9-1.9H72.04C70.99,112.37,70.13,113.22,70.13,114.27z\"\n />\n </g>\n </svg>\n <span class=\"product-documents__info\">\n SNAP-CUTANA™ K-MetStat Panel Shell Script</span\n >\n </a>\n </div>\n <div class=\"product-documents\">\n <a\n href=\"/content/documents/SNAP-CUTANA_K-MetStat_Panel_Analysis.xlsx\"\n target=\"_blank\"\n class=\"product-documents__link\"\n >\n <svg\n version=\"1.1\"\n id=\"Layer_1\"\n xmlns=\"http://www.w3.org/2000/svg\"\n xmlns:xlink=\"http://www.w3.org/1999/xlink\"\n x=\"0px\"\n y=\"0px\"\n viewBox=\"0 0 228 240\"\n style=\"enable-background: new 0 0 228 240\"\n xml:space=\"preserve\"\n class=\"product-documents__icon\"\n >\n <g>\n <path\n class=\"product-documents__svg-excel\"\n d=\"M193.17,69.88l-46.83-46.83c-1.31-1.31-3.07-2.05-4.92-2.05H48.96C45.11,21,42,24.11,42,27.96v181.07\n c0,3.85,3.11,6.96,6.96,6.96h139.29c3.85,0,6.96-3.11,6.96-6.96V74.82C195.21,72.97,194.47,71.19,193.17,69.88L193.17,69.88z\n M179.15,78.02h-40.96V37.06L179.15,78.02z M179.54,200.33H57.67V36.67h65.73v47.01c0,5.05,4.09,9.14,9.14,9.14h47.01V200.33z\n M119.06,133.32l-13.45-22.29c-0.48-0.78-2.24-1.24-2.24-1.24h-8.33c-0.5,0-0.98,0.13-1.39,0.41c-1.21,0.76-1.58,2.36-0.8,3.6\n l17.85,28.28l-18.08,28.8c-0.76,1.22-0.39,2.83,0.84,3.6c0.41,0.26,0.89,0.39,1.37,0.39h7.48c0,0,1.74-0.26,2.22-1.02l13.65-22.09\n l13.56,22.07c0.48,0.78,2.22,1.26,2.22,1.26h8.18c0.5,0,0.98-0.15,1.42-0.42c1.22-0.79,1.57-2.4,0.79-3.63l-18.35-28.48\n l18.63-28.94c0.78-1.23,0.42-2.85-0.81-3.63c-0.42-0.27-0.9-0.41-1.4-0.41h-7.79c0,0-1.76,0.7-2.24,1.48L119.06,133.32z\"\n />\n </g>\n </svg>\n <span class=\"product-documents__info\"\n >SNAP-CUTANA™ K-MetStat Panel Analysis</span\n >\n </a>\n </div>\n </div>\n </div>\n </div>\n </div>\n</div>\n","tags":[],"warranty":"","price":{"without_tax":{"formatted":"$295.00","value":295,"currency":"USD"},"tax_label":"Sales Tax"},"detail_messages":"","availability":"","page_title":"SNAP-CUTANA K-MetStat Spike-In Control for CUT&RUN / CUT&Tag","cart_url":"https://www.epicypher.com/cart.php","max_purchase_quantity":0,"mpn":null,"upc":null,"options":[],"related_products":[{"id":694,"sku":null,"name":"CUTANA™ pAG-MNase for ChIC/CUT&RUN 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","image":{"data":"https://cdn11.bigcommerce.com/s-y9o92/images/stencil/{:size}/products/764/750/2020_CUTTag_icon_RGB_Andy_L__93238.1592491196.png?c=2","alt":"CUTANA™ pAG-Tn5 for CUT&Tag"},"images":[{"data":"https://cdn11.bigcommerce.com/s-y9o92/images/stencil/{:size}/products/764/750/2020_CUTTag_icon_RGB_Andy_L__93238.1592491196.png?c=2","alt":"CUTANA™ pAG-Tn5 for CUT&Tag"}],"date_added":"17th Jun 2020","pre_order":false,"show_cart_action":true,"has_options":true,"stock_level":null,"low_stock_level":null,"qty_in_cart":0,"custom_fields":[{"id":1005,"name":"Internal Comment","value":"extra boxes in bottom shelf of Venom"}],"num_reviews":null,"weight":{"formatted":"0.01 LBS","value":0.01},"demo":false,"price":{"without_tax":{"currency":"USD","formatted":"$765.00","value":765},"price_range":{"min":{"without_tax":{"currency":"USD","formatted":"$765.00","value":765},"tax_label":"Sales Tax"},"max":{"without_tax":{"currency":"USD","formatted":"$2,970.00","value":2970},"tax_label":"Sales 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2020","pre_order":false,"show_cart_action":true,"has_options":false,"stock_level":null,"low_stock_level":null,"qty_in_cart":0,"custom_fields":[{"id":588,"name":"Pack size","value":"48 Reactions"}],"num_reviews":null,"weight":{"formatted":"0.01 LBS","value":0.01},"demo":false,"add_to_cart_url":"https://www.epicypher.com/cart.php?action=add&product_id=772","price":{"without_tax":{"currency":"USD","formatted":"$945.00","value":945},"tax_label":"Sales Tax"},"add_to_wishlist_url":"/wishlist.php?action=add&product_id=772"},{"id":986,"sku":null,"name":"CUTANA™ CUT&Tag Kit","url":"https://www.epicypher.com/products/epigenetics-kits-and-reagents/cutana-cut-and-tag-kit","availability":"","rating":null,"brand":{"name":null},"category":["Antibodies/CUTANA™ CUT&Tag Antibodies","Epigenetics Kits and Reagents/CUTANA™ CUT&Tag Assays"],"summary":"\n \n \n \n Description\n \n \n The CUTANA™ CUT&Tag Kit offers a comprehensive so","image":{"data":"https://cdn11.bigcommerce.com/s-y9o92/images/stencil/{:size}/products/986/1117/Save_15_with_code_CTK15__15909.1678741803.png?c=2","alt":"CUTANA™ CUT&Tag Kit"},"images":[{"data":"https://cdn11.bigcommerce.com/s-y9o92/images/stencil/{:size}/products/986/1117/Save_15_with_code_CTK15__15909.1678741803.png?c=2","alt":"CUTANA™ CUT&Tag Kit"},{"data":"https://cdn11.bigcommerce.com/s-y9o92/images/stencil/{:size}/products/986/1116/Epicypher_CUTANA_CUTTag_Box__75004.1678458194.png?c=2","alt":"CUTANA™ CUT&Tag Kit"}],"date_added":"8th Mar 2023","pre_order":false,"show_cart_action":true,"has_options":true,"stock_level":null,"low_stock_level":null,"qty_in_cart":0,"custom_fields":[{"id":1219,"name":"Pack Size","value":"48 Reactions"}],"num_reviews":null,"weight":{"formatted":"0.01 LBS","value":0.01},"demo":false,"price":{"without_tax":{"currency":"USD","formatted":"$2,695.00","value":2695},"tax_label":"Sales 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Description
The SNAP-CUTANA K-MetStat Panel of spike-in controls for CUT&RUN and CUT&Tag offers an all-in-one solution to determine antibody specificity for histone posttranslational modifications (PTMs), monitor assay success, and normalize data for quantitative chromatin mapping. The panel contains designer nucleosomes (dNucs) representing 16 different K-methyl PTM states: mono-, di-, and trimethylation at H3K4, H3K9, H3K27, H3K36, & H4K20, as well as unmodified control (Figure 1). Each PTM is represented by two unique DNA-barcoded templates (A and B, for an internal technical replicate). Each dNuc is individually conjugated to paramagnetic beads and pooled into a single panel for convenient one-step spike-in to CUT&RUN and CUT&Tag experiments. The panel is added to samples alongside ConA-immobilized cells prior to the addition of an antibody targeting a histone lysine methylation state or IgG negative control (see Application Notes and Table 1). pAG-MNase-mediated release or pAG-Tn5-mediated tagmentation of genomic chromatin and the barcoded nucleosomes is dependent on the specificity of the antibody used. After sequencing, the relative read count of each spike-in nucleosome barcode provides a quantitative metric of on- vs. off-target recovery (Figures 2 and 3) as well as quantitative sample normalization, thereby gauging experimental success, guiding troubleshooting efforts, and enabling reliable cross-sample comparisons.
The SNAP-CUTANA Spike-in User Guide - everything you need to know in one place
Validation Data
Figure 1: Schematic of SNAP-CUTANA Spike-in
Controls
The K-MetStat Panel controls for 15 widely studied methyl
states (see Description) and an unmodified control (me0),
with each of the 16 octamers wrapped with two uniquely
barcoded DNA templates (A and B). Each 250 bp DNA template
contains a 123 bp 601 nucleosome positioning sequence (gray)
[1], a unique 22 bp DNA-barcode (white; 32 barcodes total),
and a 5’ biotin-TEG. The 5’ and 3’ linkers (blue) are
compatible with cleavage by pAG-MNase (EpiCypher
14-1048,
15-1016) during CUT&RUN as well as tagmentation by pAG-Tn5
(EpiCypher
15-1017
) during CUT&Tag. The dNucs are individually pre-conjugated
to paramagnetic beads and pooled for convenient use.
Figure 2: Panel Identity Profiling by CUT&RUN
CUT&RUN was performed on 500k fresh K-562 cells utilizing
best in class identified antibodies to each PTM represented
in the SNAP-CUTANA K-MetStat Panel. Prior to antibody
addition, 2 µL of the control panel was spiked into each
sample. CUT&RUN sequencing reads were aligned to the unique
DNA barcodes in the panel and normalized to either on-target
(anti-K-methyl PTM) or total counts (IgG, Input). Antibody
enrichment of the expected spike-in nucleosome target (red)
confirmed the identity and integrity of each member of the
panel. CUT&RUN with IgG control antibody showed no
preferential enrichment for any particular nucleosome
(blue), as expected. Direct sequencing of the panel (Input)
indicated balanced pooling of each nucleosome. While
EpiCypher has not yet identified an H4K20me1 antibody with a
preferred specificity profile in CUT&RUN, the clone shown
has sufficient PTM preference to confirm the integrity of
the barcoded nucleosome.
Note: While most antibodies used in these
experiments show good target specificity in our
standard CUT&RUN / CUT&Tag conditions,
changes to antibody lots and experimental variables (cell
number, cell type, experimental treatment, salt
concentration, etc.) can have adverse effects on antibody
specificity.
Single-point antibody validation under optimal conditions
is not a suitable substitute for controlled experiments
with spike-ins.
EpiCypher recommends using SNAP spike-in controls to monitor
experimental success and accuracy in every possible
reaction.
Figure 3: Panel Identity Profiling by CUT&Tag
CUT&Tag was performed on 100k fresh K-562 cells utilizing
best in class identified antibodies to each PTM represented
in the SNAP-CUTANA K-MetStat Panel. Prior to antibody
addition, 2 µL of the control panel was spiked into each
sample. CUT&Tag sequencing reads were aligned to the unique
DNA barcodes in the panel and normalized to either on-target
(anti-K-methyl PTM) or total counts (IgG). Antibody
enrichment of the expected spike-in nucleosome target (red)
confirmed the identity and integrity of each member of the
panel. CUT&Tag with IgG control antibody showed no
preferential enrichment for any particular nucleosome
(blue), as expected.
Note: While most antibodies used in these
experiments show good target specificity in our
standard CUT&RUN / CUT&Tag conditions,
changes to antibody lots and experimental variables (cell
number, cell type, experimental treatment, salt
concentration, etc.) can have adverse effects on antibody
specificity.
Single-point antibody validation under optimal conditions
is not a suitable substitute for controlled experiments
with spike-ins.
EpiCypher recommends using SNAP spike-in controls to monitor
experimental success and accuracy in every possible
reaction.
Table 1: Recommended SNAP-CUTANA Spike-In Dilution for
CUT&RUN / CUT&Tag Samples of Varying Starting Cell
Number
Figure 4: DNA Gel Data
Representative images for SNAP-CUTANA K-MetStat nucleosomes
resolved by native PAGE and stained with ethidium bromide to
confirm intact nucleosome assembly with minimal free DNA.
Lane 1: Free 250 bp DNA used in nucleosome assembly (100
ng). Lane 2: Intact nucleosomes (400 ng). Comparable
experiments were performed for the entire SNAP-CUTANA
K-MetStat Panel.
Figure 5: Protein Gel Data
Representative Coomassie stained PAGE of SNAP-CUTANA
K-MetStat dNucs (1 µg each) demonstrates the purity of
histones in the preparation. Sizes of molecular weight
markers and positions of the core histones (H2A, H2B, H3 and
H4) are indicated. Comparable experiments were performed for
the entire SNAP-CUTANA K-MetStat Panel.
Technical Information
Application Notes
See the most recent EpiCypher CUT&RUN or CUT&Tag protocols for detailed information on workflow integration, expected results, data analysis, and troubleshooting. In Brief:
Product Use: Use the SNAP-CUTANA K-MetStat Panel for control reactions containing positive (e.g. H3K4me3) and negative (IgG) antibodies, as well as samples with an antibody to any of the 15 lysine methyl states in the K-MetStat Panel. Just before antibody addition, spike in 2 μL per 500k cells for CUT&RUN and 2 μL per 100k cells for CUT&Tag. If using less than the standard number of cells, decrease the amount of SNAP-CUTANA spike-in linearly by preparing a “working stock” dilution of the panel in the appropriate buffer, made fresh the day of use (Table 1). Adjust spike-in volume as needed; aiming for the spike-ins to comprise ~1% of the total sequencing reads. Expect higher for low abundance targets / negative controls (IgG; ~10-20%) and lower for high abundance targets (H3K27me3; 0.1-1%). Table 1 gives recommended dilution amounts for varying numbers of starting cells, but optimization may be required for user-specific conditions.
Data Analysis: Perform paired end sequencing for a minimum of 50 bases. The Widom 601 DNA and DNA barcodes are distinct from human, mouse, fly, and yeast genomes such that they can be readily distinguished from sample chromatin. A shell script (.sh file extension) for spike-in alignment and an excel template for heatmap generation are available in the Documents & Resources section below. The shell script can be opened with any basic text editor program and contains detailed instructions hashed (#) at the beginning of the document. Make sure to copy and paste the R1 & R2 echo loop so there is a set for each sample being analyzed.