The ABI 4800 is a tandem time-of-flight MS/MS system and is the workhorse of the MBCF workflow. The 4800 offers high accuracy intact mass analysis of all types of samples as well as ms/ms capabilities for fragmentation analysis of peptides and other bio-molecules It’s high accuracy, (<10ppm), speed, and easy sample prep make analyzing your sample so simple, researchers can be trained for walk-up use.
Generally, we start processing samples as soon as they arrive. For intact MW determinations this means you usually get data back within 2 hours of receipt of sample.
For Digest and MassFingerprinting allow 2 days for processing.
For LC-MS/MS results allow for additional 2 days.
Intact Mass Analysis
Concentration: Optimally, 10uM. Samples at 1uM work if they are clean and very low buffer and salt content.
Buffer: Samples presented in water or other volatile solvents are best. If your sample is not compatible with those requirements, then use a low salt buffer (< 50mM). Try to avoid phosphate buffers as they quench the signal on the MALDI. Avoid DMSO or glycerol if possible. If not, keep percentage of glycerol/DMSO to <5%.
Volume: 1-10ul. More is better.
Containers: Eppendorf type tubes, with your initials, sample name and date clearly on tube.
Samples for digest and protein identification:
Amount: Optimally, >500 fmol or >50 ng. We can detect as low as 5 ng of protein but the coverage is not great, and the identification is less likely to work. For post-translation modification searches, it is important to give at least 4-5 pmol of sample.
Stain: Coomasie, Sypro, or other mass spec compatible silver stain. Stay away from silver stain containing gluteraldehydes. Do not over-stain gel.
Controls: Please provide blank portion of gel. For your benefit provide a known, not pre-stained, standard. Controls are not charged.
Containers: Place gel slice in methanol rinsed 1.5ml Eppendorf tube.(No need for additional liquid). Clearly label all tubes with your initials, sample name and date.
1st sample, +blank,
|Digest and MALDI