How does Helicos see single molecule fluorescence?

[Nathan McCorkle]

This video says they take sheared ssDNA, add polyA tail, hybridize
about 1 molecule per square micron on a surface coated with polyT,
then add polymerase and a single species of fluorescent nucleotide,
rinse, illuminate with laser light and scan the surface with a camera,
cleave fluorophore, then repeat with a different species of nucleotide
and more polymerase.

http://www.helicosbio.com/Portals/0/Videos/tSMS-How_It_Works.flv

I wonder if they're using a single pixel camera with a PMT backend?

I wonder if I could replicate with just 1 pixel (1 molecule only), I
could use this to provide feedback for closed-loop DNA synthesis
operations.

--
Nathan McCorkle
Rochester Institute of Technology
College of Science, Biotechnology/Bioinformatics

[Nathan McCorkle]

Thanks John!

I imagine the CCD is back-thinned and cooled, and from the size of the
Helicos cabinet (and the video showing the CCD samples hybridization
area in sections), I wonder if they're stepping the optics/wafer
around like a microchip mask aligner... with servos and encoders for
rough positioning, and interferometers for fine positioning (unless
they ensure their stepper motors are repeatable enough, through high
holding torque, if they're using steppers)

Seems like a PMT would do the job for a single reaction center!

On Fri, Aug 24, 2012 at 3:23 PM, singlemolecule
<thompso...@gmail.com> wrote:
> If you want more information on what the optical/camera system looks like
> (as well as more detail on other aspects), go to
> http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2954431/ and look at figure 2.
>
> S

> --
> You received this message because you are subscribed to the Google Groups
> "DIYbio" group.
> To post to this group, send email to diy...@googlegroups.com.
> To unsubscribe from this group, send email to
> diybio+un...@googlegroups.com.
> To view this discussion on the web visit
> https://groups.google.com/d/msg/diybio/-/w5qlL3GV-cEJ.
> For more options, visit https://groups.google.com/groups/opt_out.

[Nathan McCorkle]

Here's a patent of theirs:
http://www.google.com/patents?id=TfEAAgAAEBAJ

[singlemolecule]

Helicos is the only system that can generate usable reads directly from RNA and it is vastly superior to any other current system for quantitative accuracy in applications like ChIP Seq and RNA Seq where read length and accuracy are less important than number of reads and lack of amplification.  Despite the technical advantages, commercial uptake has been extremely limited for a variety of reasons and this is not improving.  A quick look at the stock price will confirm that.

S

On Saturday, September 1, 2012 7:06:29 AM UTC-4, hrusty wrote:

Hello, I have been a follower of Helicos for a few years now. My question is to your technical expertise. Do you see this platform competitive in the current market or is it a league of its own because tSMS and DRS and lack of amplification making a direct threat to the staus quo?

Thanks for your time

[John Griessen]

On 09/04/2012 10:34 AM, singlemolecule wrote:
> commercial uptake has been extremely limited for a variety of reasons

What reasons?

[singlemolecule]

One reason was instrument price.  When this became coupled with concerns about long-term viability, it created a downward spiral.