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Differential scanning fluorimetry based assessments of the thermal and kinetic
stability of peptide-MHC complexes
#MMPMID26906089
Hellman LM
; Yin L
; Wang Y
; Blevins SJ
; Riley TP
; Belden OS
; Spear TT
; Nishimura MI
; Stern LJ
; Baker BM
J Immunol Methods
2016[May]; 432
(?): 95-101
PMID26906089
show ga
Measurements of thermal stability by circular dichroism (CD) spectroscopy have
been widely used to assess the binding of peptides to MHC proteins, particularly
within the structural immunology community. Although thermal stability assays
offer advantages over other approaches such as IC50 measurements, CD-based
stability measurements are hindered by large sample requirements and low
throughput. Here we demonstrate that an alternative approach based on
differential scanning fluorimetry (DSF) yields results comparable to those based
on CD for both class I and class II complexes. As they require much less sample,
DSF-based measurements reduce demands on protein production strategies and are
amenable for high throughput studies. DSF can thus not only replace CD as a means
to assess peptide/MHC thermal stability, but can complement other peptide-MHC
binding assays used in screening, epitope discovery, and vaccine design. Due to
the physical process probed, DSF can also uncover complexities not observed with
other techniques. Lastly, we show that DSF can also be used to assess peptide/MHC
kinetic stability, allowing for a single experimental setup to probe both binding
equilibria and kinetics.
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