Thursday, June 3, 2010

Intellicyt HyperCyt Impressions

Well, the HyperCyt has come and gone and to sum up my thoughts in a word I can only say, Wow.  When it comes to speed, this thing is really fast.  I tried to put a bunch of different things through the sampler to try and get it to fail, but it just didn't; big stuff, clumpy stuff, small stuff, dyes, you name it.
Now, let me put things into perspective for you a bit.  We run everything in tubes manually, so to run through 96 samples I've been able to achieve a throughput rate of about 40 minutes (collecting a couple of thousand cells per sample).  My target for a sampler was to get an equivalent amount and quality of data in less than 20 minutes per plate.  By many accounts, this is NOT high throughput.  The representative from Intellicyt actually sort of chuckled at me when i was explaining this.  His idea of high throughput was collecting 384 well plates in about 6 or 7 minutes.  I certainly can appreciate the utility of this unit for something like that, but my standards of data quality are probably a bit more strict when compared to a "screening" assay.
What I was really impressed with was the ability to precisely control every aspect of the sampling process such that you can go really fast, or pretty slow if you wanted to.  What I am showing here is a comparison of the same exact plate run at a "high" speed (~11 minutes per 96 well plate) and at a "low" speed (~27 minutes per 96 well plate).  It is a simple Annexin V FITC vs. 7-AAD cytotoxicity assay, that is comparing the affects of a drug on the cells.  As you can see, there is no appreciable difference in the two, and they both are the same as when I ran these samples manually in a tube.

Other things I ran were two big cell lines (one GFP, one RFP) mixed together to look at the potential increase in coincidence events (double positives) when going at high rates.  When I ran the sample manually in a tube, I saw about 0.5% double positive cells.  When I ran them through the HyperCyt at the higher flow rate, that double positive population shot up to 1.6%, and when I ran it at the slower flow rate, it was about 1%.  The reason this happens, as I'm told, is that when the unit sends the bolus of cells into the cytometer, the cells tend to accumulate at the leading edge of the bolus, so you get a high concentration of cells right in the beginning of collection, thus leading to higher coincidence.  In fact when I looked at the formation of coincident events versus time of a single collected sample I saw exactly that, all the double positives were detected in the 1st second or so of collection, then after that, it leveled out to the 0.5% seen while running the samples manually in a tube.
One other thing I was concerned with was the residual dyes left over when rinsing between samples.  To test this, I ran alternating samples of PI stained CENs and DAPI stained CENs.  I first ran it with no wash to show that the probe itself may be transferring dye from well to well (which it did).  Then I ran the samples with a wash in between collection and then there was no increase in PI staining of the DAPI CENs and no increase in DAPI staining of the PI CENs.  This, along with multiple other tests run with beads put me at ease as to the capabilities of this instrument as a multi-well plate sampler, not just a high throughput screen tool.
So, will I get one?  Likely, but possibly in the form of a Accuri/HyperCyt combo ala what was displayed at the CYTO 2010 meeting.  This is a new product from Intellicyt, which they're calling the HTFC (High Throughput Flow Cytometer).  Here's a link to the info on that (warning, this is a link to a pdf, so it will automagically start downloading to your computer; not to worry though, it's safe).

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