Chronological List of Experiments
 

4 new slides #36-41, 5 Mar 2003
All stained for NuMA and with dapi.

slide36   - B1067 T6 slide of frozen mammary tissue
slide37   - B1069 T6 slide of frozen mammary tissue
slide38 -   S1 EGFR 3D drip Day 10
slide39 -   T4 cells 3D drip Day 10
slide40 -   S1 60 monolayer Day 4
slide41 -   T4 cells monolayer Day 3
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Subject:   Sent slides today
Date:  Tue, 4 Mar 2003 14:31:41 -0500
From:  cmbator@purdue.edu
To:   David Knowles <dwknowles@lbl.gov>
CC:   Sophie Lelievre <lelievre@purdue.edu>

Hi David,

This is to let you know that I did get the slides packaged and sent out to you by FedEx today.  They should arrive tomorrow (Wednesday).  The FedEx tracking number is 828101858685.

There are 6 slides this time.
* B1067 T6 slide of frozen mammary tissue stained with NuMA primary antibody coded P-5.
* B1069 T6 slide of frozen mammary tissue stained with NuMA primary antibody coded B1C11.
* S1 EGFR 3D drip Day 10 wells 1 & 2 stained with NuMA primary antibody coded B1C11 and wells 3 & 4 stained with NuMA primary antibody coded P-35.
* T4 cells 3D drip Day 10 wells 1 & 2 stained with NuMA primary antibody coded B1C11 and wells 3 & 4 stained with NuMA primary antibody coded P-35.
* S1 60 monolayer Day 4 wells 1 & 2 stained with NuMA primary antibody coded B1C11 and wells 3 & 4 stained with NuMA primary antibody coded P-35.
* T4 cells monolayer Day 3 wells 1 & 2 stained with NuMA primary antibody coded B1C11 and wells 3 & 4 stained with NuMA primary antibody coded P-35.

Good Luck,
Carol Bator Kelly
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2Ap03
Conversation with Sophie:
1) because NuMA staining requires Triton extraction, it is difficult to remove acini from the Matrigel. Cell tend to clump together.
2) In slides 36-41 & 42-47, Sophie has sent S1-EGFR cells cultured in 3D for 10 days. S1 cells cultured in EGFR are unable to properly growth arrest and thus do not differenciate. Thus they have the characteristic of diplasure but are not tumeragenic. When imaging these cultures it will be important to choose thoes cells in large (ugly) clusters. By eye, NuMA distribution in S1-EGFR looks like differenciated S1.
3) 3D drip is a method of 3D culture.
4) 2 points in the first paper. Firstly, it is important to use 3D cultures to study nuclear distribution because we have shown that NuMA distributes very differently in 3D verses 2D cultures. Secondly, our ability to discern non-malignant from malignant even at early growth.
5) The image Sophie sent, NuMAforDavid2.jpg, shows NuMA distribution in a non-differentiated T4 nucleus, a differentiated S1 3D day10 nucleus and an S1-EGFR 3D day10 nucleus. The T4 nucleus shows diffuse staining. The arrows in the differentiated nucleus show NuMA rings at the perimeter and the arrow head points to a tripple punctate spot.