Guideline for DNA Sample Collection for Epiontis’ Epigenetic qPCR Immunomonitoring Service DNA Isolation To isolate genomic DNA from cells, tissue or whole blood the "DNeasy Blood & Tissue Kit" from Qiagen (Cat. No. 69504) is recommended. A pdf version of the „DNeasy Blood & Tissue Handbook“ is available online at www1.qiagen.com. Please refer to the handbook for purification protocols and sample starting amounts. For a high yield DNA isolation, please start with about 100 µl blood or a similar amount of cells as suggested in the protocol, but replace the 100 µl elution buffer with 50 µl of elution buffer, and use this amount to re-elute the column 3 times. We recommend in general sending an amount of 10 µg DNA per sample in a volume of 50 µl or less. This relatively high amount allows a re-measurement of your samples (at no additional cost) in case a QC parameter for a plate run is not met or if a sample has an unusually low target cell content. If sample amounts are limited, please refer to the table on the second page of this guideline for minimum DNA inputs that should be provided. Shipping to Epiontis (Germany) • To avoid any customs delays, it is recommended to specify samples in the cover or consignment letter as follows: genomic DNA of human origin, non-toxic, non-hazardous, non-infectious, for research purpose only, commercial value: 1 US $ / 1 Euro. • DNA samples can be sent at 4°C using cool packs, shipment on dry ice is not necessary. • Use clearly labelled safe lock tubes additionally sealed with parafilm to avoid leakage. • Please ship samples to: Epiontis GmbH Attention Julia Oppatt Rudower Chaussee 29 12489 Berlin Germany • Please notify Epiontis prior to shipment to check for German holidays and provide the carrier’s tracking number as soon as the samples have been shipped (see email below). Epiontis will monitor the progress of shipment and confirm sample receipt. Version No.: 05 Replaces Version No.: 04 QMF 508-1e Page: 1 of 2 Minimum DNA inputs Cell Type T cells Regulatory T cells (Treg) B cells Th17 cells T Helper cells Cytotoxic T cells Neutrophil Granulocytes CCR-6 positive cells Granulolysin positive cells NK cells Monocytes TFH cells Typical Level in Blood of Healthy Donors [%] DNA input amount 31.3% 1.7% 4.1% 2.6% 15.1% 9.1% 55.6% 7.3% 2.6% 2.8% 10.2% 4.2% 570 ng 2.6 µg 780 ng 3.8 µg 300 ng 480 ng 540 ng 2.5 µg 2.1 µg 2.6 µg 300 ng 620 ng In more than 98% of measurements the minimum input will ensure measurements in the validated assay range with standard precision if the sample contains at least half of the typical target cell level observed with the epigenetic assay in blood of healthy donors. Contact Julia Oppatt Manager Analytical Laboratory [email protected] Version No.: 05 Replaces Version No.: 04 Epiontis GmbH Rudower Chaussee 29 12489 Berlin. Germany QMF 508-1e Fax: +49 30 6392 3476 www.epiontis.com Page: 2 of 2