RE: SPAM-LOW: Re: ISAC tries to forget San Diego and Harry Crissman?
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From: FloCyte Training Institue <flocyte@[EMAIL PROTECTED]
>
Date: Tue Jun 17 2008 - 12:24:05 EDT
And isn't there a council member missing? Tim Bushnell??
Sue
-----Original Message-----
From: J. Paul Robinson [mailto:jpr@[EMAIL PROTECTED]
Tuesday, June 10, 2008 3:01 PM
To: cyto-inbox
Subject: SPAM-LOW: Re: ISAC tries to forget San Diego and Harry
Crissman?
Well sorry about that Kevin - it was on the original list for
sure...and
it was on the copy we handed out in Quebec...and it was up in the
ISAC
booth at Budapest....but you are right its missing in the current
handbook...my apologies - and I think grounds for firing
me....oh...too
late for that I guess now!!
regards
paul
is in the C. Kevin Becker wrote:
> Hi All,
>
>
>
> I was thumbing through the new ISAC member****p handbook, looking at the
> color pages on the different ISAC meetings and there seems to be an ISAC
> meeting missing. ISAC XXI, 2002, San Diego, Harry Crissman, President.
>
>
>
> Is this discrepancy just my book or everyone's? I thought we had a
> pretty good time at the banquet with the Mardels and doesn't anybody
> remember riding the wave at the PFS booth?
>
>
>
> Regards,
>
>
>
> Kevin
>
>
>
> C. Kevin Becker
>
> Phoenix Flow Systems, Inc.
>
> 6790 Top Gun St. #1
>
> San Diego, CA 92121
>
> 858 453-5095
>
> fax 858 453-2117
>
> www.phoenixflow.com
>
>
>
> -----Original Message-----
> *From:* Arthur Roberts [mailto:robertar@[EMAIL PROTECTED]
> *Sent:* Friday, May 30, 2008 9:33 AM
> *To:* Cytometry Mailing List
> *Subject:* SUMMARY: PI viability dye during sorting
>
>
>
> Dear Flow-ers,
> Thank you for all your useful replies. They ran quite the gamut, with
> all sorts of interesting suggestions. I am posting a synopsis here.
>
> The original question: I was wondering about the risks in using PI in a
> stream-in-air sorter. Since PI is a DNA intercalater, and therefore,
> could have nasty effects on the operator's DNA if it were inhaled as an
> aerosol, is there a way to use it in sorting that will minimize
> exposure? I would wash the cells before sorting, but it seems that the
> PI intensity would drop off quickly after wa****ng, since the binding is
> reversible.
>
> Thanks for any insight you might have.
>
> - Art
>
> Arthur Roberts
> Robert Wood Johnson Medical School
> Dept. of Molec. Genetics, Microbiol & Immunol
> SRB rm 113
> 675 Hoes Lane
> Piscataway, NJ 08854
> phone: 732-235-4502
>
> REPLIES:
>
> Actually the PI binding is quite stable; we see little loss in
> fluorescence over a couple of hours.
>
> I would bet the amount of PI aerosolized during pipetting far exceeds
> what you would get from a sorter.
> ______________________________
>
> As an alternative you may want to use the food dye that is used to
> discard dead and damaged sperm during sperm sorting for ***
> predetermination. A 0.002% solution of this dye, FD&C #40, (Johnson and
> Welch. Theriogenology 1999;52:1323-1321; Garner and Seidel. Reproduction
> 2002;124:733-743) is added to Hoechst 33342 stained sperm. This dye
> identifies the dead and damaged sperm so that they can be discarded in
> the waste fluid at the time of sorting. This system is being used
> commercially and we have had no indication that it damages the sorted
> living sperm (Garner and Seidel, Theriogenology, 2008;69:886-895)
>
> The red dye quenches the Hoechst 33342 and with the dual detectors ends
> up as a smeared population in the lower quadrant of the bivariate plot
> of UV emission detectors.
>
>
>
> The FD & C #40 powder is procured from the Warner Jenkinson Co., 2526
> Baldwin St., P.O. Box 14538, St.Louis, MO 63178-0538 .
>
> One dissolves 5 g of the FD&C #40 in 100 mL nanopure water to make a 5%
> stock solution to use for staining samples.
>
>
>
> ______________________________________
>
>
>
> We use PI all the time, it is at a rather low concentration and
> so I have never worried about it. we have a stock that is at 1mg/ml
> which we dilute into a working
> solution 50 fold less concentrated, then add that to 200 to 400=B5l of
> cells. This works out to a final concentration of 0.5-1=B5g/ml if I did
> my math right.
> _______________
>
> Yes, it is called aerosol containment devices, no sort operator should
> be caught without it. If your instrument doesn=92t have one then you
> should be seriously consider using a Personal Protective device. What do
> you do when you sort other live material? I think I would be more
> concerned about aerosol inhalation of other biohazardous materials from
> live cells. Kevin Holmes did a very nice evaluation of aerosol
> generation from an Aria cell sorter characterizing both size and
> quantity of aerosols generated under a variety of conditions and showed
> a good percentage are just the right size for inhalation into the lung.
> He presented this at ISAC and will be presenting it at the upcoming
> Chesapeake Cytometry meeting next Monday in Rockville, MD. Sorting
> without some type of aerosol containment practice is at best risky.
> _________________________
>
> A correctly set up stream-in-air sorter should pose no risk at all.
> I ran a MoFlo for 3 1/2 years, when PI was the viability & cell cycle
> dye of choice.
> I agree aerosols are generated if using mixed populations such as blood,
> especially at the maximum event rate for sorting. However, the amount
> of PI in your sample that is being liberated as an aerosol throughout
> the sort is minute, it would be an interesting exercise to even try and
> measure it.
> My exposure was simply limited by a perspex box that was fitted to the
> front of the MoFlo during a sort. Any basic air extraction (with Hepa
> filters) would also be worthwhile. My current sorter (an Aria - which
> has a sealed sort chamber) came with a very neat extraction system that
> is never used.
>
> Invitrogen are selling some very nice dyes for viability/cell
> cycle/live-dead now that are supposed to be considerably safer than PI.
> __________________________
>
> Last advice that I had was that PI hazard is mainly from the powder
> form. If the operator only works with PI solution then the hazard is
> greatly reduced. I still would not handle the PI solution without
> gloves, though, and I feel sympathy for the poor person who has to make
> up the solution from powder! I did some tests a while ago as to the
> washout effects on PI, and found no significant leaching in the cells I
> was using.
>
> _______________________________
>
> PI would be the last of my worries in live samples.
> ________________________________
>
>
>
--
J. Paul Robinson
SVM Professor of Cytomics
Professor of Immunopharmacology & Biomedical Engineering
Director, Purdue University Cytometry Laboratories
Past-President, International Society for Analytical Cytology
(now International Society for Advancement of Cytometry)
Purdue University Cytometry Laboratories
Bindley Bioscience Center
1203 West State Street
Discovery Park, Purdue University
West Lafayette, IN 47907-2057
Ph (765) 494 0757; Fax (765) 494 0517
email: jpr@[EMAIL PROTECTED]
ISAC - www.isac-net.org
Change lives today - www.cytometryforlife.org
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6:29 PM
Received on Wed Jun 18 14:58:00 2008
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