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 msms 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 walkup 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