Educational guides and overviews on research peptides, what they are and how they're studied in the laboratory. For research use only.
BLI reports the optical thickness of a layer at a fibre tip as a wavelength shift in nanometres; every rate constant after that is inference. What the dip-and-read format changes about loading, referencing, mass transport and rebinding compared with SPR, and why fast-off peptide interactions defeat it.
Read moreHow an MST trace is built from a temperature jump, directed migration and re-equilibration, and how label chemistry, detergent, capillary adsorption and aggregation get into the number. Written for deciding what an MST Kd licenses next to ITC and SPR.
Read moreHow amide backbone exchange chemistry, the EX1 and EX2 regimes, quench conditions and back-exchange set the limits of an HDX-MS dataset. Includes why a peptide that takes up less deuterium on ligand binding is not thereby a binding site.
Read moreA crosslink is a proximity measurement with an upper bound, not an interaction. This guide covers crosslinker chemistry and spacer arms, the quadratic search problem, false discovery rate estimation at spectrum, residue-pair and protein-pair level, and the Calpha-Calpha bounds that turn a hit into a restraint.
Read moreHow a Gq calcium transient is generated, read and mis-read: indicator chemistry and buffering, dye loading and anion-transport blockers, peak versus area under the curve, promiscuous and chimeric Galpha subunits, and receptor reserve. Written for deciding what a calcium EC50 does and does not license.
Read moreAspartyl residues cyclize onto their own backbone under repeated piperidine treatment, and the ring-opened products are mostly invisible to intact mass. This guide covers the mechanism, the sequence dependence, the product distribution, and the four independent ways to suppress it.
Read moreWhy substoichiometric phosphorylation forces an enrichment step, how IMAC and TiO2 differ in selectivity, recovery and multiply-phosphorylated bias, and what a site-localization score and its false localization rate actually license you to claim.
Read moreHow the choice of connectivity, the ring-closure site and the linear precursor's conformation decide whether a macrocyclization gives one product or a mixture. Covers the epimerization-versus-oligomerization activation dilemma, effective molarity, turn inducers, on-resin versus solution closure, and what the entropy argument does not license.
Read moreA working account of copper-catalysed and strain-promoted azide-alkyne cycloaddition on peptides: catalyst and ligand stoichiometry, the oxidative damage the ligand suppresses, cyclooctyne rate and lipophilicity trade-offs, azide and alkyne building blocks, and the residues where each chemistry stops being orthogonal.
Read moreHow orthogonal synthetase/tRNA pairs are selected, why amber suppression is a race against release factor 1, and what residue-specific replacement in an auxotroph can and cannot give you. Includes why a band at the right molecular weight is not evidence of incorporation.
Read moreHow N- and O-linked attachment chemistry, anomeric control and glycan acid/base lability constrain Fmoc glycopeptide assembly, and what oxonium ions, ETD/EThcD and exoglycosidase digestion each license you to conclude about a glycosylation site.
Read moreHow peptide electrophoretic mobility, wall-generated electroosmotic flow, coatings and Joule heating together set what a capillary electrophoresis method can resolve. Includes why charge-difference isomers and D/L pairs need a handle other than hydrophobicity.
Read moreHow drift-tube, travelling-wave and trapped ion mobility turn an arrival time into a collision cross section, what a CCS distribution licenses you to conclude about gas-phase conformer populations, and where that conclusion stops short of solution structure.
Read moreA TR-FRET reading says two labels came close, not how close. What the lanthanide donor and the time gate actually suppress, what the two-wavelength ratio is licensed to correct, why the r^-6 dependence makes this proximity rather than distance, and the hook that leaves one sandwich reading ambiguous.
Read moreHow chiral derivatisation turns amino acid enantiomers into separable diastereomers, why total hydrolysis both enables and corrupts the measurement, and what a D-percentage actually licenses you to claim.
Read moreOxazolidine and thiazolidine rings turn serine, threonine and cysteine into temporary prolines that kink the backbone and come off again in TFA. How the ring is built, where to place it, why it is sold as a dipeptide, and what Hmb and Dmb cost you when the sequence offers no hydroxyl or thiol.
Read moreHow to get a peptide alpha-thioester into your hands when Fmoc chemistry makes direct preparation inefficient: Boc assembly, safety-catch and N-acylurea linkers, and the intramolecular N-to-S acyl shift devices. Covers thiol leaving-group reactivity, additive exchange in the ligation buffer, and the scoped evidence on epimerisation and hydrolysis.
Read moreSortase A cleaves LPXTG, requires Cys184, which points to a thioacyl intermediate, and hands the acyl group to an oligoglycine amine. The product is still a substrate, and that single fact governs enzyme loading, nucleophile excess and the strategies for driving the reaction to completion.
Read moreAn absorbance at 280 nm measures tryptophan, tyrosine and cystine, then converts them into peptide by an assumed coefficient. Where that coefficient comes from, how folding, denaturants and neighbouring residues move it, what the 205 and 214 nm alternatives cost in matrix sensitivity, and how a UV number is anchored to a mass-balance number.
Read moreA stability-indicating method is the claim that your analyte separates from everything it can turn into, and the way that claim gets tested is by making the degradants on purpose. The target degradation window and why over-stressing generates products the material will never make, what peak purity by diode array and by mass spectrometry each establish, mass balance and what a deficit means, and identifying degradants rather than merely resolving them.
Read moreWhere the solvents in a lyophilised peptide come from, why the diluent decides the answer more than the column does, when standard addition is the calibration the matrix forces on you, and what the number cannot tell you about water or counterion.
Read moreSize-exclusion spin columns, dialysis, tangential flow and reversed-phase SPE can each lose a different part of the sample. What the molecular-weight cutoff actually rates, why recovery collapses at low input, what carried-over salt does at the ion source, and when the step changes the peptide itself.
Read moreAn MD ensemble is a force field's prediction, not a measurement. Known force-field biases for disordered and helical peptides, what implicit solvent trades away, the gap between accessible simulation time and the process you care about, what a free-energy surface from replica exchange, metadynamics or umbrella sampling does and does not establish, and how far agreement with NMR, order-parameter and CD data actually gets you.
Read moreReplacing the sugar-phosphate backbone with N-(2-aminoethyl)glycine buys a neutral strand that binds harder and discriminates better, and costs you phosphoramidite chemistry, aqueous solubility on long purine-rich sequences, and membrane permeability. What the Boc and Fmoc routes actually deliver, why purine-rich chains aggregate on resin, how mismatch position and identity move Tm, and what a melting curve does and does not license.
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