What Receptor Binding Kinetics & SPR got wrong, from 26 theses

Tried and failed

surface plasmon resonance and quartz crystal microbalance applied to protein binding and unbinding kinetics. Reason: severe protein aggregation and irreversible non-specific deposition formed thick layers on the sensor surface

Biophysical Dynamics Of Rgs-Lov Proteins As Systems For Light-Induced Membrane Recruitment · Penn · PhD

Tried and failed

affinity enrichment without signal amplification applied to secreted glycoproteins in serum-containing media. Outcome: no signal. Reason: Non-specific binding of high-abundance background proteins overwhelmed target glycoprotein detection.

MS-Based Chemical Proteomics Studies of Extracellular Glycoproteins: Identification, Quantification, and Dynamics · Georgia Tech · PhD

Tried and failed

liposome sedimentation assay for membrane protein recruitment applied to protein-protein complex recruitment to lipid membranes. Outcome: no signal. Reason: proteins exhibited high non-specific intrinsic binding to synthetic liposomes without target complex present

STRUCTURAL AND BIOCHEMICAL MECHANISMS OF THE ACTIVATION OF ARF1 AT THE GOLGI · Cornell · PhD

Tried and failed

SPME-based measurement of specific protein binding affinity applied to long-chain hydrophobic surfactant-protein interactions. Outcome: no signal. Reason: Non-specific sorption dominated, masking any saturable high-affinity specific binding sites on the protein

Toxic Effects and Exposure of Per- and Polyfluoroalkyl Substances in Cell-based Bioassays · Tübingen · PhD

Lost to a baseline

Hydrophobic long-chain alkanethiol SAM-functionalized Au surfaces exhibited significantly stronger non-selective binding (NSB) of BSA compared to polystyrene 96-well ELISA plates and hydrophilic dextran-functionalized CM5 chips

Methods to Improve the Stability and Selectivity of Electronic Biosensors · Georgia Tech · PhD

Considered and rejected

Considered and rejected: Rejected poly-L-lysine (PLL) slide coating due to prominent non-specific probe binding background

Single-cell imaging assay for rapid antimicrobial resistance testing · Oxford · Master's

Considered and rejected

Considered and rejected: Ceased testing synthetic peptide binders (RRGW, WGRR, Fc-III-4C) because binding signal was below or equivalent to non-specific background binding to ELISA plates.

Collagen Anchoring Agonist Antibodies for Cancer Immunotherapy · MIT · PhD

Tried and failed

screening cyclic peptide libraries at low ionic strength applied to protein-protein interaction inhibitor discovery. Reason: low ionic strength causes non-specific electrostatic binding and high false-positive rates

Discovery of macrocyclic inhibitors of challenging protein-protein interactions · EPFL · PhD

Tried and failed

ELISA screening of bacterial expression clones applied to single-domain antibody selection. Reason: Screening selected non-specific plastic-binding clones rather than target-specific binders, generating false positives.

Targeting anti-PD-1 and anti-PD-L1 single-domain antibodies to the tumour microenvironment via matrix metalloproteinase activity · Imperial · PhD

Tried and failed

baculovirus particle ELISA using recommended coating dilution applied to antibody specificity screening. Reason: Recommended dilution caused false positive non-specific binding from direct antibody binding to the microplate

Specificity and other Biophysical Properties to Guide Developability of Agonistic Bispecific Antibodies · Harvard · Master's

Considered and rejected

Considered and rejected: Rejected direct physical adsorption antibody immobilization due to excessive non-specific binding and non-uniform patchiness across gold wires.

Microfluidics Sensor to Detect Water Salinity and Bacteria Using Optical and Electrical Methods · York University · Master's

Tried and failed

amine coupling surface plasmon resonance immobilisation applied to recombinant protein on sensor chip. Reason: protein failed to immobilise under standard low pH sodium acetate buffer conditions

Targeting anti-PD-1 and anti-PD-L1 single-domain antibodies to the tumour microenvironment via matrix metalloproteinase activity · Imperial · PhD

Tried and failed

high surface density ligand immobilization applied to biosensor nanoparticle binding detection. Outcome: worse than baseline. Reason: excessive surface packing density caused severe steric hindrance that impeded analyte binding kinetics and affinity

DIRECT DETECTION OF EXTRACELLAR VESICLE TRANSMEMBRANE PROTEIN – ANTIBODY INTERACTIONS USING SURFACE PLASMON RESONANCE BIOSENSORS · Cornell · PhD

Considered and rejected

Considered and rejected: Rejected amine coupling of galectins directly to SPR chips because covalent immobilization disrupted oligomerization and caused loss of binding activity.

Defining the mechanism of galectin-3-mediated TGF-β1 activation and its role in lung fibrosis · Nottingham · PhD

Considered and rejected

Considered and rejected: Rejected long-linker biotinylation reagents (20,000 g/mol) despite superior MACS capture efficiency due to steric hindrance attenuating scFv-antigen binding.

Developing an antibody discovery platform for targeting membrane proteins via cell-cell screening · Johns Hopkins · PhD

Tried and failed

responsive polymer shell coating for plasmonic biosensing applied to protein detection via LSPR shifts. Outcome: no signal. Reason: Target protein binding caused no concentration-dependent localized surface plasmon resonance wavelength shifts across various polymerizations

Impact of synthesis technique and compositon of polymer coatings for semiselective gold nanoshell-based biosensors · UT Austin · PhD

Tried and failed

direct protein physisorption onto sensor surface applied to optical biosensing transducers. Outcome: no signal. Reason: insufficient surface binding density or low refractive index contrast prevents measurable optical resonance shifts

All-Dielectric Nanophotonic via Glass Fluid Instabilities · EPFL · PhD

Considered and rejected

Considered and rejected: Rejected surface plasmon resonance (SPR), ITC, and MST for determining ion binding affinity due to molecular weight mismatch or redox metal instability, proposing DSF instead

Copper A Novel Regulator Of Perk · Penn · PhD

Considered and rejected

Considered and rejected: Rejected using isothermal titration calorimetry (ITC) and surface plasmon resonance (SPR) for the benzophenanthridinone class (968) due to high protein concentration requirements where GAC forms dimers/tetramers rather than binding monomers.

THE ALLOSTERIC REGULATION OF GLUTAMINASE ENZYMES: DRIVING GLUTAMINE CATABOLISM IN CANCER CELLS · Cornell · PhD

Considered and rejected

Considered and rejected: Standard binding assays (QCM, SPR, floatation assays, fluorimetry, calorimetry) were rejected in favor of high-throughput AR-SHS because QCM loses sensitivity below nanogram protein weights, SPR requires kinetic modeling limiting sensitivity to the micromolar range, fluorimetry requires labeling that perturbs native binding affinity, and calorimetry lacks binding-event specificity.

Second harmonic scattering of water in a biological context · EPFL · PhD

Considered and rejected

Considered and rejected: Rejected NMR titration and isothermal titration calorimetry (ITC) for measuring aqueous nanohoop-fullerene binding affinities due to low sensitivity (>10^6 M^-1) and precipitation artifacts.

Functionalized Carbon Nanohoops: Nitrogen-Doped Partial Belts, Macrocyclic Ligands, and The Inherent Strain That Affects Their Chemical Properties · University of Oregon · PhD

Considered and rejected

Considered and rejected: Rejected surface plasmon resonance (SPR) for determining LO1 and LO1Fab binding kinetics because the large size and heterogeneity of MDA-LDL yielded unreliable and inaccurate data; adopted microfluidic diffusional sizing instead.

Anti-oxidised low-density lipoprotein antibodies and antibody fragment targeted nanoparticles for the translational study of atherosclerosis · Imperial · PhD

Tried and failed

surface plasmon resonance binding affinity modeling applied to fragment library screening on structured RNA. Outcome: did not converge. Reason: complex multi-site binding and solubility limits prevented reaching surface saturation needed for kinetic fitting

Can In-Silico Computational Virtual Screening of Fragment Libraries Model In-Vitro Biophysical Screening Methods with Similar Results to Identify Novel RNA-Binding Chemical Matter · Harvard · Master's

Tried and failed

surface plasmon resonance kinetic fitting applied to fast-exchanging protein-protein interactions. Reason: association and dissociation rates were too fast to be resolved within instrument temporal limits

CHARACTERIZING THE STRUCTURE AND DYNAMICS OF ADAPTIVE IMMUNE PROTEINS TO INFORM IMMUNOTHERAPY DESIGN · Penn · PhD

Considered and rejected

Considered and rejected: Discontinued SPR screening for small-molecule KD determination after recurring sensorgram distortions and non-saturating kinetics.

The development and evaluation of PET imaging agents targeting PD-L1 · Imperial · PhD

Lost to a baseline

In QCM flow mode, fLNP mass loading on alumina surfaces was lower than on silica and gold surfaces due to electrostatic binding causing mass transfer limitations.

Development of nanoporous packaging for improving stability and pulmonary delivery of vaccine formulations · Imperial · Doctorate

Lost to a baseline

WGM lysozyme adsorption exhibited significantly slower binding kinetics than Biacore SPR and Q-Sense QCM due to microfluidic flow rate and dead volume limitations.

Opracowanie i zastosowanie mikroczujników fluorescencyjnych do nowej, bezznacznikowej metody detekcji wykorzystujacej zjawisko optycznego rezonansu Whispering Gallery Mode (WGM) Development and application of fluorescent microsensors for new label-free detection method based on Low-Q Whispering Gallery Modes · Adam Mickiewicz University · PhD

Left open

Assess antibody affinity and somatically hypermutated clones using surface plasmon resonance and B-cell sequencing instead of ELISA titers. Blocker: Requires a wet lab, animal/patient samples, surface plasmon resonance apparatus, and B-cell sequencing facilities.

Mesoporous Silica Rods Scaffolds for the generation of adaptive immune responses · Harvard · PhD

Left open

Identify additional hits from antibody libraries and characterize full IgG binding affinity using surface plasmon resonance. Blocker: Requires wet lab facilities, surface plasmon resonance equipment, and physical antibody libraries/samples

ENGINEERING THE HUMORAL RESPONSE TO GENERATE ANTIGEN-SPECIFIC ANTIBODIES · Cornell · PhD

Left open

Validate binding affinity of ambiguous or fluorescence-quenching MST hit compounds against G9a using surface plasmon resonance assays. Blocker: Requires a wet lab, surface plasmon resonance instrumentation, purified recombinant G9a protein, and physical compound samples

Development and evaluation of potential co-factor competitive inhibitors for the histone lysine methyltransferase G9a · Imperial · PhD

Left open

Optimize and repeat surface plasmon resonance (SPR) binding kinetics measurements between knuckle peptides and BMP receptors. Blocker: Requires wet lab access, an SPR instrument, synthesized knuckle peptides, and purified BMP receptors

Design considerations of BMP-2 mimicking peptide immobilized scaffolds for use in bone tissue engineering · Imperial · PhD

Left open

Perform direct binding validation and biophysical characterization of non-aromatic bacterial quorum sensing molecule DPD with AhR. Blocker: Requires wet lab assays and biophysical characterization apparatus (e.g., purified AhR protein and DPD binding assays)

REGULATON OF TYPE I INTERFERON SIGNALING AND HOST DEFENSE AGANST BACTERIA BY 2,3,7,8-TETRACHLORODIBENZO-p-DIOXIN-INDUCIBLE POLY(ADP-RIBOSE) POLYMERASE · Cornell · PhD

Left open

Test ribosome binding sites of varying strengths to measure the effect of different translation rates on plasmid crosstalk in cell-free biosensing. Blocker: Requires molecular biology wet lab experiments with plasmid variants in cell-free extracts.

Towards Robust, Point-of-Care Biosensing in Cell-Free Systems · Georgia Tech · PhD

Left open

Test whether a simple cell or protein binding assay predicts loss of binding and in vivo transcytosis activity of antibody mutants. Blocker: Requires wet lab experiments with cell or protein binding assays and mutant antibodies

Development and Exploration of Immunohistochemistry as An in vitro Assay for Predicting Antibody Transcytosis in vivo · Harvard · Master's