high sensitivity reagents

Bill Telford telfordw at box-t.nih.gov
Mon Aug 21 15:35:11 EST 2000


Hi David...

You may be thinking of the PBXL fluorochromes manufactured by Martek
Biosciences and marketed by Intergen and KPL.  This fluorochrome is
actually an intact stabilized  cyanobacterial photosystem II
phycobiliprotein complex consisting of an allophycocyanin core and chains
of phycoerythrins linked to the core with phycocyanins, making it a huge
complex (several million daltons).  The thing operates as it would in vivo,
namely as a massive energy transfer complex, and should be extremely bright
as a result.  They have two - PBXL-1, which excited at 488 nm and emits
around 660 nm, and PBXL-3, which excites around 620 nm red (i.e. red HeNe
or diode laser range) and emits around 660 nm.

We tried these complexes a couple of years ago for flow and found their
large size to be problematic with regard to steric hindrance - they were as
bright as APC, but should have been much brighter given their structure.
Their massive size was undoubtedly interfering with ligand binding.  At the
time their use didn't seem to be justified given their sensitivity
equivalence to the much smaller APC complex.  The Martek group has since
generated PBXL-3 avidin conjugates with PEG spacer arms - these show much
lower levels of steric hindrance.  We now can generally get at least 5- to
20-fold enhancement in sensitivity with PBXL-3 over APC, with relatively
low backgrounds.  We now use PBXL-3-PEG-streptavidin in several systems
where we need to detect weakly expressed surface markers.

The fluorescence products group at Martek (F.C.Tom Allnutt, Mark Moss and
John Morseman) have been very helpful to us in working with their
conjugates.  Their contact info is...

Martek Biosciences Corporation
Fluorescence Diagnostics Products Group
6480 Dobbin Road
Columbia, MD 21045
phone:   (410) 740-0081
WWW:  http://www.martekbio.com

Regards,

Bill Telford

NCI MB Flow Cytometry Core Laboratory
DCS-NCI-NIH

At 12:35 PM 8/18/00 -0400, you wrote:
>
>Good day to all.  Sometime ago I received an add for antibody-fluorescent
>conjugate that delivered on the order of 100 molecules of dye per binding
>event.
>)Alright, I don't know if was really 100, but it was a lot.)  It was a big
>phycobiloprotein construct as I recall.  Does this ring a bell with anyone?
>I'm trying to detect very low expression of a surface antigen and thought this
>might be the way to go.  The add must have gotten lost during my recent
>move.  I
>hope someone can help because it's kinda hard to search for something if you
>don't know what it's called!
>
>David McFarland
>SmithKline Beecham Pharmaceuticals
>
>





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