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Homogeneous sample preparation for automated high throughput analysis with matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry.
Rapid Commun Mass Spectrom. 1999; 13(5):315-22.RC

Abstract

This work presents a simple method for obtaining homogeneous sample surfaces in matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOFMS) for the automated analysis of peptides and proteins. The sample preparation method is based on applying the sample/matrix mixture onto a pre-deposited highly diluted matrix spot. The pre-deposited crystals act as seeds for the new sample containing crystals which become much smaller in size and more evenly distributed than with conventional methods. This 'seed-layer' method was developed, optimised and compared with the dried-droplet method using peptides and proteins in the 1000-20,000 Da range. The seed-layer method increases the surface homogeneity, spot to spot reproducibility and sample washability as compared with the commonly used dried-droplet method. This methodology is applicable to alpha-cyanohydroxycinnamic acid, sinapinic acid and ferulic acid, which all form homogeneous crystal surfaces. Within-spot variation and between-spot variation was investigated using statistics at a 95% confidence level (n = 36). The statistical values were generated from more than 5000 data points collected from 500 spectra. More than 90% of the sample locations results in high intensity spectra with relatively low standard deviations (RSDs). Typically obtained data showed an RSD of 19-35% within a sample spot as well as in-between spots for proteins, and an RSD of < or = 50% for peptides. Linear calibration curves were obtained within one order of magnitude using internal calibration with a point-RSD of 3% (n = 10). The sample homogeneity allows mass spectra (average of 16 laser shots) to be obtained on each individual sample within 15 sec, whereby a 100 spot target plate can be run in 25 min. High density target plates using the seed-layer method were prepared by spotting approximately 100 picoliter droplets onto the target, resulting in sample spots < or = 500 microns in diameter using a flow-through piezo-electric micro-dispenser. By using this automated sample preparation step lower standard deviations are obtained in comparison to manually prepared samples.

Authors+Show Affiliations

Onnerfjord P
Department of Analytical Chemistry, Lund University, Sweden.
Ekström S
No affiliation info available
Bergquist J
No affiliation info available
Nilsson J
No affiliation info available
Laurell T
No affiliation info available
Marko-Varga G
No affiliation info available

MeSH

Coumaric AcidsCrystallizationInsulinMicroscopy, Electron, ScanningPeptidesProteinsSpectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

Pub Type(s)

Journal Article

Language

eng

PubMed ID

10209870

Citation

Onnerfjord, P, et al. "Homogeneous Sample Preparation for Automated High Throughput Analysis With Matrix-assisted Laser Desorption/ionisation Time-of-flight Mass Spectrometry." Rapid Communications in Mass Spectrometry : RCM, vol. 13, no. 5, 1999, pp. 315-22.
Onnerfjord P, Ekström S, Bergquist J, et al. Homogeneous sample preparation for automated high throughput analysis with matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry. Rapid Commun Mass Spectrom. 1999;13(5):315-22.
Onnerfjord, P., Ekström, S., Bergquist, J., Nilsson, J., Laurell, T., & Marko-Varga, G. (1999). Homogeneous sample preparation for automated high throughput analysis with matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry. Rapid Communications in Mass Spectrometry : RCM, 13(5), 315-22.
Onnerfjord P, et al. Homogeneous Sample Preparation for Automated High Throughput Analysis With Matrix-assisted Laser Desorption/ionisation Time-of-flight Mass Spectrometry. Rapid Commun Mass Spectrom. 1999;13(5):315-22. PubMed PMID: 10209870.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Homogeneous sample preparation for automated high throughput analysis with matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry. AU - Onnerfjord,P, AU - Ekström,S, AU - Bergquist,J, AU - Nilsson,J, AU - Laurell,T, AU - Marko-Varga,G, PY - 1999/4/21/pubmed PY - 2000/6/20/medline PY - 1999/4/21/entrez SP - 315 EP - 22 JF - Rapid communications in mass spectrometry : RCM JO - Rapid Commun Mass Spectrom VL - 13 IS - 5 N2 - This work presents a simple method for obtaining homogeneous sample surfaces in matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOFMS) for the automated analysis of peptides and proteins. The sample preparation method is based on applying the sample/matrix mixture onto a pre-deposited highly diluted matrix spot. The pre-deposited crystals act as seeds for the new sample containing crystals which become much smaller in size and more evenly distributed than with conventional methods. This 'seed-layer' method was developed, optimised and compared with the dried-droplet method using peptides and proteins in the 1000-20,000 Da range. The seed-layer method increases the surface homogeneity, spot to spot reproducibility and sample washability as compared with the commonly used dried-droplet method. This methodology is applicable to alpha-cyanohydroxycinnamic acid, sinapinic acid and ferulic acid, which all form homogeneous crystal surfaces. Within-spot variation and between-spot variation was investigated using statistics at a 95% confidence level (n = 36). The statistical values were generated from more than 5000 data points collected from 500 spectra. More than 90% of the sample locations results in high intensity spectra with relatively low standard deviations (RSDs). Typically obtained data showed an RSD of 19-35% within a sample spot as well as in-between spots for proteins, and an RSD of < or = 50% for peptides. Linear calibration curves were obtained within one order of magnitude using internal calibration with a point-RSD of 3% (n = 10). The sample homogeneity allows mass spectra (average of 16 laser shots) to be obtained on each individual sample within 15 sec, whereby a 100 spot target plate can be run in 25 min. High density target plates using the seed-layer method were prepared by spotting approximately 100 picoliter droplets onto the target, resulting in sample spots < or = 500 microns in diameter using a flow-through piezo-electric micro-dispenser. By using this automated sample preparation step lower standard deviations are obtained in comparison to manually prepared samples. SN - 0951-4198 UR - https://www.unboundmedicine.com/medline/citation/10209870/Homogeneous_sample_preparation_for_automated_high_throughput_analysis_with_matrix_assisted_laser_desorption/ionisation_time_of_flight_mass_spectrometry_ L2 - https://doi.org/10.1002/(SICI)1097-0231(19990315)13:5&lt;315::AID-RCM483&gt;3.0.CO;2-C DB - PRIME DP - Unbound Medicine ER -
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