sorting large particles

Witherspoon, Sam sw11527 at
Mon Jul 16 10:04:13 EST 2001

Hi again folks,
	Eric, I did not mention this product at first because I did not have
the info at my fingertips. (also did not think you would be interested in
having to buy anything but....)

There is a company which makes a "large bead compatible" instrument
(originally made to sort nematodes) which has been investigated here and
seems to work.

Union Biometrica
US Headquarters
35 Medford Street, Suite 213
Somerville, Massachusetts 02143
Tel: 617-591-1211
Fax: 617-591-8388

Their website has changed since I last visited but there is still an
overview of their system.

I had not realized how old this particular technology was until I found this
Automated sorting and dispensing of C. elegans to wells of microtiter plates
CD Johnson1, R Clover1, B Reardon1, PB Krauledat2, AA Ferrante2, WP Hansen2
1	Axys Pharmaceuticals, NemaPharm Group, South San Francisco, CA.
2	Union Biometrica, Inc., Somerville, MA.
For generating a large number of separate populations of worms, liquid
cultures in the wells of microtiter plates use space more efficiently than
do standard cultures on agar plates. At NemaPharm, we routinely grow worms
in microtiter plates for a wide variety of procedures including both forward
and reverse genetics experiments as well as for high-throughput chemical
screening. To automate distribution of worms into microtiter plate wells, we
have constructed a machine that efficiently sorts and dispenses live
The mechanism used for measuring nematodes is based, in principle, on the
nematode counting machine built by Lou Byerly, Randy Cassada and Dick
Russell in the early 1970's (1). Nematodes are suspended in liquid and
passed though a narrow nozzle into the center of a rapidly flowing sheath
current. The resultant shear forces straighten the animals and orient them
parallel to the direction of flow. The straightened animals are then passed
at high speed (~1 m/sec) through an orthogonal sheet of laser light where,
in the current model, an in-line detector measures the attenuation of the
laser light and an attached computer calculates the length of the nematode
based upon time-of-flight through the beam. Below the detector, the
worm-containing stream is deflected to waste by a air jet controlled by the
computer that shuts off momentarily to deliver a selected animal to a well
(distribution volume is 0.25 - 1 µl per worm).
The current sorter/dispenser is capable of distributing 10 worms to the
wells of 96-well plates in 2 minutes with a precision of ±0.7 worms.
Distribution to 384 and 1536 well plates is also feasible with the current
model. Machines with more sophisticated detecting and measuring capabilities
are planned for the future.

(1) Byerly, et. al. (1975) Rev Sci. Instrum., 46:517-522

Sam Witherspoon
sw11527 at
High Throughput Biology
*				 Tel. 919-483-3078
GlaxoSmithKline R&D		 Page 919-857-7768
5 Moore Dr.			 Fax  919-483-0585
RTP, NC  27709

> -----Original Message-----
> From: Nealley, Eric W Mr USAMRICD [SMTP:Eric.Nealley at]
> Sent: Wednesday, July 11, 2001 10:11 AM
> To:	Cytometry Mailing List
> Subject:	sorting large particles
> Our group has been tasked to sort large diameter beads (180 um) which have
> gone through a process to attach peptides.  We suggested that a new
> library
> of smaller peptide labeled beads be used, but for now the group wants to
> try
> and make use of the larger beads until a new library can be created using
> small beads.	We have a FACStar Plus equipped with a 400 um nozzle (with
> turbo sort option) and we have tried to align/standardize/calibrate with
> little success. We have followed the BD macrosort procedure (lower sheath
> psi and lower ddf) but again with little success. The only bead that I
> currently have for alignment are flow check beads which are ~10 um (I
> assume
> at this point that 10 um beads are not the correct ones to use with the
> large nozzle).  I have ordered 100 um beads from FCSC.
> -  Has anyone sorted using a 400 um nozzle?
> -  Has anyone successfully sorted ~180 um beads before?
> -  How do we align/calibrate/standardize the instrument?
> Thank you.
> Eric Nealley
> Biologist, Pharmacology Division
> Aberdeen Proving Ground, Maryland

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