Bioaffinity Screening with a Rapid and Sample-Efficient Autosampler for Native Electrospray Ionization Mass Spectrometry.
Anal Chem
; 93(39): 13342-13350, 2021 10 05.
Article
em En
| MEDLINE
| ID: mdl-34546705
ABSTRACT
Fast and efficient handling of ligands and biological targets are required in bioaffinity screening based on native electrospray ionization mass spectrometry (ESI-MS). We use a prototype microfluidic autosampler, called the "gap sampler", to sequentially mix and electrospray individual small molecule ligands together with a target protein and compare the screening results with data from thermal shift assay and surface plasmon resonance. In a first round, all three techniques were used for a screening of 110 ligands against bovine carbonic anhydrase II, which resulted in five mutual hits and some false positives with ESI-MS presumably due to the high ligand concentration or interferences from dimethyl sulfoxide. In a second round, 33 compounds were screened in lower concentrations and in a less complex matrix, resulting in only true positives with ESI-MS. Within a cycle time of 30 s, dissociation constants were determined within an order of magnitude accuracy consuming only 5 pmol of ligand and less than 15 pmol of protein per screened compound. In a third round, dissociation constants of five compounds were accurately determined in a titration experiment. Thus, the gap sampler can rapidly and efficiently be used for high-throughput screening.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Pesquisa
/
Espectrometria de Massas por Ionização por Electrospray
Tipo de estudo:
Diagnostic_studies
/
Screening_studies
Limite:
Animals
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article