Antibodies and reagents:
... LPS (List Biological Laboratories), ... were used to stimulate BMDCs.
Author did not specify which List Labs LPS product was utilized in their research. List Labs provides the following LPS products: https://www.listlabs.com/product-information/lipopolysaccharides/
Enzyme-Linked ImmunoSorbent Assay (ELISA):
96 well microtiter plates (EIR/RIA plate, Costar, Lowell, MA) were coated with 100 μl rPA (List labs, Campbell, CA) at 1 μg/ml in PBS overnight (o/n) at 4°C. Plates were subsequently blocked with 200 μl 1% BSA (in PBS containing 0.05% Tween 20, PBS-T) for two hours at 4°C. ...
TLR Stimulation and Cytokine Measurement:
For ELISA, day 5 BM-derived macrophages were plated in a 96-well tissue plate (5 × 104 cells per well) overnight. Titrations of TLR stimuli were added for 16 h as follows: Salmonella minnesota R595 LPS (List Biological Laboratories), ...
Materials and Methods:
SNAPtide® substrates (Products #521 and #523), Unquenched Calibration Peptide for SNAPtide® 521 (Prod #528), BoNT/A LC (Product #610A), and BoNT/A holotoxin (Prod #130A) are products of List Biological Laboratories, Inc. ...
Methods - The above materials were used in the following (refer to poster for details):
SNAPtide®substrate (Product # 520) and Botulinum Neurotoxin Type A (Product #130A) are products of List Biological Laboratories.
Summary of Results: The His6-SV2c-His6 was purified in a soluble form and linked to magnetic nickel coated beads. Various buffer conditions and binding conditions were tested to optimize the binding and limit the non specific binding of toxin to the beads. We are able to detect cleavage of SNAPtide®after exposure to as little as 5pg of BoNT/A.
Conclusions: Our study shows the ability to purify a soluble form of the SV2c BoNT/A binding domain and its utility in the capture and detection of BoNT/A. This is a key step in establishing an in vitropotency assay. This assay can also be used to screen for inhibitors that interfere in the binding of BoNT/A to the SV2c protein receptor.