The present specification discloses SNAP-25 compositions, methods of making α-SNAP-25 antibodies that bind an epitope comprising a carboxyl-terminus at the P.sub.1 residue from the BoNT/A cleavage site scissile bond from a SNAP-25 cleavage product, α-SNAP-25 antibodies that bind an epitope comprising a carboxyl-terminus at the P.sub.1 residue from the BoNT/A cleavage site scissile bond from a SNAP-25 cleavage product, methods of detecting BoNT/A activity, and methods of detecting neutralizing α-BoNT/A antibodies.

Provided herein is a cell-free assay device, sometimes comprising a lipid bilayer and an endopeptidase assay component, for characterizing a pore forming protein. In some embodiments provided herein is an apparatus comprising a pressure system for characterizing an interaction. Also, provided herein are methods for using a cell-free assay device to characterize a pore forming protein and/or a test substance.

The invention provides methods for analysing a sample of immunoglobulins to determine the amount of IgG aggregates present in the sample.

Methods for detecting biomarkers of inflammation, infection, and/or bacterial activity in dairy production, which indicate issues with the milk itself or issues related to the health of the cow. The methods generally comprise contacting a milk sample with a nanoplatform assembly to create an assay solution, and detecting spectral changes in the assay solution that are triggered by enzymatic activity (when present) in the sample. The nanoplatform assembly comprises a first particle, a second particle, and a linkage therebetween, wherein the linkage comprises a protease consensus sequence (the sequence of amino acids cleaved by the protease), or an ester linkage (cleaved by a protease or lipase). A plurality of second particles can also be linked to the first particle. Test strips are also described, which undergo a visual color change in the presence of the target enzyme in the milk sample.

The present disclosure relates generally to peptide reporter constructs of SNAP-25, which are useful in determining the activity of botulinum toxins, and methods of using the same.

The present invention makes it possible to simply, rapidly, and with high sensitivity detect periodontopathic bacteria at room temperature and in under ten minutes (approximately 2-8 minutes). The arginine-specific peptidase activity of three strains of periodontopathic bacteria, i.e., Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia, are analyzed at room temperature, in under ten minutes, and with high sensitivity. A first compound that has an anti-oxidizing effect and/or a second compound that protects SH groups (mercapto groups) and has a disulfide bond-cleaving effect are present during analysis of a sample. The first compound is at least one among L-ascorbic acid, L-cysteine hydrochloride, and glutathione, and the second compound is at least one among DTT, thioglycolic acid, thioglycerol, mercaptoethanol, and TCEP.

Compositions and methods for improved cell-based methods of characterizing botulinum neurotoxins are provided. Cells utilized in these methods include a reporting construct that is cleaved following uptake and processing of botulinum neurotoxin by the cell, resulting in proteolysis of the portion of the reporting construct that is released following cleavage. The released portion includes a fluorophore and amino acid substitutions or sequences that enhance the rate of proteolysis. A pair of reporting constructs can be utilized in which one member of the pair is modified to resist cleavage by the botulinum neurotoxin while co-localizing with the remaining member of the pair.

Disclosed herein is a botulinum neurotoxin (BoNT) polypeptide with a modified receptor binding domain (HC) having one or more amino acid mutations that modify the binding of the BoNT to the receptor. Specific mutations and combinations of mutations are also disclosed. Isolated modified HC, polypeptides comprising the modified HC, chimeric molecules, pharmaceutical compositions, and methods of making and using the same are also disclosed. Methods of identifying additional such modified receptor binding domains, are further disclosed.

Biosensors including a nucleic acid encoding a PcaU protein, a PobR protein, a CatM protein, a PcaR protein, or a TphR protein are provided. In some examples, the biosensors include a promoter regulated by the sensed molecule operably linked to a reporter gene. The biosensors may be included in a vector or in cells including one or more of the biosensors or vectors. Modified chorismate pyruvate lyase (UbiC) and modified paraoxonase (PON1) proteins including one or more amino acid substitutions are provided. Finally, methods of selecting biocatalysts with increased activity including transforming a library of cells expressing a biosensor with one or more nucleic acids encoding one or more mutations in a gene involved in a biosynthesis pathway, determining activity of the reporter protein; and selecting a cell with increased reporter protein activity as expressing a biocatalyst with increased activity are provided.

The present specification discloses SNAP-25 compositions, methods of making α-SNAP-25 antibodies that bind an epitope comprising a carboxyl-terminus at the P.sub.1 residue from the BoNT/A cleavage site scissile bond from a SNAP-25 cleavage product, α-SNAP-25 antibodies that bind an epitope comprising a carboxyl-terminus at the P.sub.1 residue from the BoNT/A cleavage site scissile bond from a SNAP-25 cleavage product, methods of detecting BoNT/A activity, and methods of detecting neutralizing α-BoNT/A antibodies.