Ultrasonic multiprobe as a new tool to overcome the bottleneck of throughput in workflows for protein identification relaying on ultrasonic energy.
We studied in this work the performance of the new ultrasonic multiprobe in terms of throughput, handling and robustness. The study was conducted using the multiprobe to speed two different proteomics workflows. The "classic" method relaying on overnight protein digestion (12h), was used as the standard procedure. This work clearly shows the importance of testing variables such as ultrasonic amplitude and ultrasonic time when adapting an ultrasonic-based treatment to a new ultrasonic device. The results here presented also shown and confirm the advantage of speed up sample treatment workflows with the aid of ultrasonic energy in combination with a 96-well plate. The methods compared were similar in terms of robustness, but the desalting free method was the fastest, requiring only 2 min/sample for completion. In addition it was also the simplest in terms of handling, since no desalting step was needed. The following standard proteins were successfully identified using the methods studied: bovine serum albumin, alpha-lactalbumin, ovalbumin, carbonic anhydrase, fructose-bisphosphate aldolase A, catalase, chymotrypsinogen A. As case study, the identification of the protein Split-Soret cytochrome c from D. desulfuricans ATCC 27774 was carried out.
Santos
Hugo M
HM
REQUIMTE, Departamento de Química, Centro de Química Fina e Biotecnologia, CQFB, Faculdade de Ciências e Tecnologia, FCT, Universidade Nova de Lisboa, UNL, Quinta da Torre, 2829-516 Monte de Caparica, Portugal.
Carreira
Ricardo
R
Diniz
Mario S
MS
Rivas
M G
MG
Lodeiro
Carlos
C
Moura
José J G
JJ
Capelo
José L
JL
D000818:Animals; D002417:Cattle; D045304:Cytochromes c / Q000032:analysis; D057166:High-Throughput Screening Assays / Q000379:methods*; D010449:Peptide Mapping; D011506:Proteins / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013997:Time Factors; D014465:Ultrasonics; D014508:Urea / Q000737:chemistry
D011506:Proteins; D014508:Urea; D045304:Cytochromes c