Date: Sat, 04 Jan 1997 17:19:49 -0700
From: DBrune@asu.edu (Dan Brune)
Subject: Re: MALDI
To: Recipients of ABRF List <abrf@aecom.yu.edu>
Michail,
In looking at the messages on the ABRF bulletin board after the
Christmas break, I noticed yours about about which instruments people would
recommend for MALDI-TOF mass spectromety. I think a better question might
be to ask people which features they like or do not like about the
instruments that they are using. Instrumentation in this area is changing
rapidly, and many of the instruments that were sold even a few years ago
may lack features that are fairly standard on some of the newer
instruments. Consequently recommendations people make about the
instruments that they have may not be the best guide for what kind of
instrument you should get now.
Having said that, I shall now try to answer your question as
rephrased by me. I tend to agree with Murray Hackett's comments that these
discussions should be held openly, and therefore shall venture to state my
comments on this subject in public. This also has the advantage that if I
say something that other people disagree with, thay can respond and I may
learn something from the response.
I have been using a Vestec instrument that has several features
that I consider desirable. One of these is a video camera that looks into
the sample compartment, allowing one to see (from matrix fluorescence)
where the laser is hitting the sample probe. A second uesful feature is
real time data collection and display by a 500 MHz digitizing oscilloscope.
This allows mass spectra to be observed (albeit in a signal averaged form)
as adjustments in laser intensity and sample position are being varied. It
also allows the user to look at a mass spectrum before deciding whether or
not it should be saved on the computer hard disk. A few months ago, I
recall hearing from someone at PerSeptive Biosystems (which acquired Vestec
a couple years ago) comment that they had "improved" the mass spectrometer
by getting rid of the oscilloscope. If true, this seems like a step
backward to me.
One feature that I am not particularly fond of in my instrument is
the sample handling system. Samples are placed on the ends of stainless
steel pins about 2 mm in diameter. This works well for water-soluble or
water&acetonitrile soluble samples like most proteins, where 1.5-2uL
samples can be spotted onto the pin, but samples in solvents with low
surface tension tend to run off the sides of the probe tip. PerSeptive now
uses sample plates incorporating depressions in a 10 x 10 grid into which
the samples can be spotted. I have not used this system, but to me it
seems like an improvement.
Of course, good instruments should also have high resolution and
mass accuracy. My Vestec instrument, with a resolution of about 400, was
state of the art in 1992 when we got it, but is now definitely outmoded
when compared to instruments incorporating delayed extraction. Several
instruments include this feature, which can substantially increase
resolution, although with decreasing effectiveness as the mass increases -
I recall people from PerSeptive claiming resolutions around 10,000 for
insulin at about 5700 Da, but this resolution decreased to about 850 for
carbonic anhydrase at about 29,000 Da. Delayed extraction (which may be
designated by other names by other venders - I seem to recall hearing it
also referred to as time lag focussing, for example) also subtantially
improves mass accuracy, particularly when using external standards for
calibration.
Resolution is also improved by having a reflectron (which my
instrument also lacks). My impression from looking at data generated with
instruments having reflectrons is that this is accompanied by some loss of
signal strength. Thus instruments that can be used either with or without
the reflectron would seem to be the best. A reflectron is also useful for
looking at post source ion fragmentation, and in the case of polypeptides,
this can yield sequence information.
I should add that I am not a mass spectrometrist by training, but
have been using this technique in my protein chemistry core facility at
Arizona State. Consequently, you may be able to get more expert opinions
from other people who are better trained in this area. Nevertheless, I
imagine that your uses for the instrument will be similar to mine, so I am
giving you my impressions in the hope that you will find them useful.
Best wishes,
Dan Brune
Dept. of Chemistry & Biochemistry
Arizona State University
Tempe, AZ 85287-1604
Tel: (602) 965-0795