TITLE: DNA isolation from Formalin-Fixed, Paraffin-Embedded (FFPE) material
AUTHORS: Hendrik F van Essen

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">This protocol describes the isolation of DNA from paraffin embedded tissue. The resulting DNA can be used for CGH array hybridizations. </div><div class = "text-block"><h1></h1></div><div class = "text-block">After removal of the paraffin, the tissue is lysed and the DNA is released from the cells. Then the DNA is isolated by using a silica-based column that binds the DNA.</div></div>

[STEPS]
?. [H&E staining from paraffin blocks]
Cut 3 µm (1x), 10 µm (2-3x, or more if needed depending on tumour area)
?. [H&E staining from paraffin blocks]
Mount all the section on slides with BSA 0.1%, and dry them o/n at 37°C  (to a maximum of one week)
?. [H&E staining from paraffin blocks]
Place slides in xylene
[3 times]
?. [H&E staining from paraffin blocks]
Hydrate by passing sequentially through 100%, 96%, 70% EtOH, and distilled-water
?. [H&E staining from paraffin blocks]
Wash with running tap water
?. [H&E staining from paraffin blocks]
Stain slides with eosin
?. [H&E staining from paraffin blocks]
Wash quickly in water
?. [H&E staining from paraffin blocks]
Dehydrate by passing sequentially through 70%, 96%, 100% EtOH, xylene
?. [H&E staining from paraffin blocks]
Remove slides from xylene and use DePex to mount a covering glass.
?. [H&E staining from paraffin blocks]
Let the 3 µm H&E slide be judged by the pathologist for dissection.Mark the tumor area and tumor percentage
?. [H&E staining from paraffin blocks]
Stain slides with haematoxylin (if solution is not fresh staining time can be extended, max 4 min.)
?. [DNA isolation]
Place slides in xylene 	3 x 7 min.
[3 times]
?. [DNA isolation]
Hydrate by passing sequentially through 100%, 96%, 70% EtOH, and water
?. [DNA isolation]
Stain slides with haematoxylin 	1-2 min.(if solution is not fresh staining time can be extended, max 3 min.)
?. [DNA isolation]
Wash with running tap water
?. [DNA isolation]
Place slides in distilled water
?. [DNA isolation]
Dissect the tissue from the slides (when they are still slightly wet) with a scalpel or a  needle
?. [DNA isolation]
Place the material in a clean safelock eppendorf cup
?. [DNA isolation]
Spin down in a centrifuge at full speed and get rid of the water layer
?. [DNA isolation]
Shake and invert samples (do not vortex).
?. [DNA isolation]
Add 160 µl ATL buffer
?. [DNA isolation]
Add 40 µl prot K (20 mg/mL)
?. [DNA isolation]
vortex sample Make sure all tissue is in the liquid
?. [DNA isolation]
Incubate o/n at 56°C in heat-block or waterbath and vortex regularly! Check if all tissue is digested
?. [DNA isolation]
Vortex samples and spin down
?. [DNA isolation]
Incubate at 98°C 	10 min.
?. [DNA isolation]
Spin down at 16600 rcf.	1 min.
?. [DNA isolation]
Add 200 µl AL buffer and mixvery well by vortexing!
?. [DNA isolation]
Add 200 µl Ethanol 100% and mix very well by vortexing!
?. [DNA isolation]
Incubate at RT
?. [DNA isolation]
Spin down the eppendorf cups at full speed	1 min.
?. [DNA isolation]
Transfer up to 600 µl lysate to the QIAamp MinElute Column.
?. [DNA isolation]
Spin down at 8.000 rpm
?. [DNA isolation]
Place the QIA Column in a clean 2 ml collection tube, discard the flow-throughmake sure the column tip is clear of droplets
?. [DNA isolation]
Add 500 µl AW1 buffer to the column
?. [DNA isolation]
Spin down for at 8.000 rpm and place the column in a new tube
?. [DNA isolation]
Add 500 µl AW2 buffer (QIAmp micro-kit) to the column
?. [DNA isolation]
Spin down at 8.000 rpm and place the column in a new tube
?. [DNA isolation]
Spin down at full speed to dry the membrane
?. [DNA isolation]
Place the QI column in a properly marked eppendorf cup, discard collection tube
?. [DNA isolation]
Add 20 to 50 µl (depending on sample size) of AE buffer to the column and incubate.
[room temperature]
?. [DNA isolation]
Spin down at full speed.
?. [DNA isolation]
Throw away the column, close the eppendorf cup and store the DNA at