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Differential Gene Expression Analysis of Pure Cell Populations from Frozen UterineTissue Using Laser Capture Microdissection (LCM) and cDNA Microarrays

o plain glass slides. Slides were stored at -70 oC until use.
NOTE: Slides may be stored at -70oC for up to six months.

2. Slides were stained with hematoxylin and eosin, and dehydrated per the following protocol.
1) Defrost slides briefly, 30 seconds or less.
2) Fix with 70% ethanol for 30 seconds
3) Wash with DEPC-treated distilled water for 30 seconds.
4) Stain with Mayers hematoxylin (DAKO, Cat. # S3309) for 30 seconds.
5) Wash with DEPC-treated distilled water for 30 seconds.
6) Wash with Bluing reagent (1X Scotts tap water; Sigma, Cat. # S5134) for 30 seconds.
7) Dehydrate with 70% ethanol for 30 seconds.
8) Dehydrate with 95% ethanol for 30 seconds.
9) Stain with Eosin (Sigma, Cat. # E6003) for 30 seconds.
10) Place the slides to 75% ethanol for 30 seconds.
11) Place the slides to 95% ethanol for 30 seconds.
12) Place the slides to 100% ethanol for 30 seconds.
13) Place the slides to xylene for 5 minutes.
14) Remove slides from xylene and air dry for 5 minutes.

ISOLATION OF SPECIFIC CELL POPULATION BY LCM ANALYSIS
LCM was performed using a PixCell II system (Arcturus, Cat# LCM1106) using CapSure Macro LCM caps (Arcturus, Cat# LCM0211). Pure luminal epithelium of implantation or interimplantation sites was isolated using LCM (Figure 1A), filling an entire CapSure Macro cap with cells of interest.


RNA ISOLATION AND AMPLIFICATION
1. Total cellular RNA was extracted from captured cells immediately after microdissection using the PicoPure RNA Isolation Kit (Arcturus, Cat. # KIT0202).
a) 50 l extraction buffer was pipetted into a 0.5 ml microcentrifuge tube.
b) Placed the CapSure caps carrying the captured cells onto the microcentrifuge tube and invert
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