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Page 659 out of 671

FRET Peptide Substrates for BotulinumNeurotoxins Types A, B, C, and E

Shine, N.; Endrukaite,E.; Christian, T.
Product: VAMPtide® Peptide Substrate (o-Abz/Dnp) Substratefor C. botulinum Type B Neurotoxin

Sensitive, In Vitro, Bifunctional, Potency Assay For Botulinum Neurotoxin Type A

Christian, T.; Shine, N.
Product: Botulinum Neurotoxin Type A from Clostridium botulinum

Materials:

SNAPtide® substrate #520 contains the FRET pair o-aminobenzoic acid/2,4 dinitrophenyl (o-Abz/Dnp) and SNAPtide®substrate Mca/Dnp contains (7methoxy-coumarin-4-yl)acetyl/2,4 dinitrophenyl as its FRET pair. Botulinum Neurotoxin Type A (Product #130A) and the SNAPtide®substrates are products of List Biological Laboratories.

Methods - The above materials were used in the following (refer to poster for details):

  •  ELISA assay showing specific binding of BTA to SV2c
  •  MagneGST-SV2c capture assay
  •  MagneGST-SV2c capture assay with HPLC analysis
  •  HPLC peak intensity for cleaved SNAPtide®as a function of BTA

Kinetics of Lethal Factor and Poly-D-Glutamic Acid Antigenemia during Inhalation Anthrax in Rhesus Macaques

Boyer AE, Quinn CP, Hoffmaster AR, Kozel TR, Saile E, Marston CK, Percival A, Plikaytis BD, Woolfitt AR, Gallegos M, Sabourin P, McWilliams LG, Pirkle JL, Barr JR
Product: Anthrax Lethal Factor (LF),Recombinant from B. anthracis

LF quantification:

... To achieve the necessary dynamic range and to account for high-sensitivity rapid-response sample analysis, two standard curves were prepared, one for 20 ng/ml, μl (“low volume” [LV]) and the other for 200 ng/ml, μl (“high volume” [HV]) serum volumes. In addition, three quality control samples (QCs) were prepared in human serum pools (Interstate Blood Bank, Memphis, TN) using recombinant LF (List Biological Laboratory, Campbell, CA) and validated. ...

PA antigen capture immunoassay:

Microtiter plates (Thermo Labsystems, Franklin, MA) were coated with monoclonal anti-PA antibody AVR1046 obtained by the same methodologies used for anti-LF monoclonal described previously (4) at 1.28 μg/ml in 0.01 M phosphate-buffered saline (PBS), pH 7.4. Coated plates were washed with a wash buffer consisting of PBS and 0.1% Tween 20, pH 7.4. Recombinant PA (rPA; List Biological Laboratories, Inc., Campbell, CA) was serially diluted twofold using a 1/25 or a 1/100 dilution of normal rhesus macaque serum (RMS). The rPA dilutions formed a seven-point standard curve from 0.6 to 37.5 ng/ml. ...

Author did not indicate which specific lethal factor was utilized.  List Labs provides Product #172 (Anthrax Lethal Factor (LF), Recombinant from B. anthracis) and Product #169 (Anthrax Lethal Factor (LF-A), Recombinant from B. anthracis Native Sequence).

PubMed ID: 19506008

The mesopontine rostromedial tegmental nucleus: a structure targeted by the lateral habenula that projects to the ventral tegmental area of Tsai and substantia nigra compacta

Jhou TC1, Geisler S, Marinelli M, Degarmo BA, Zahm DS
Product: Cholera Toxin B Subunit (Choleragenoid) from Vibrio cholerae in Low Salt

Tracer injections:

Several minutes after being given intraperitoneal injections of a cocktail, consisting of 45% ketamine (100mg/ml), 35% xylazine (20mg/ml) and 20% physiological saline at a dose of 0.16 ml/100g of body weight, rats were placed into a Kopf stereotaxic instrument. The skulls were exposed and small bore holes were created to allow selected brain structures to be targeted by filament-containing borosilicate glass pipettes (O.D. - 1.0 mm) pulled to tip diameters of 10–25 μm and containing the retrograde tracers Fluoro-Gold (FG; Fluorochrome, Inc., Englewood, CO; 1% in 0.1M cacodylate buffer, pH 7.4) or cholera toxin B subunit (CtB, List Biological Laboratories, Campbell, CA), ...

Author did not specify which CTB was utilized.  List Labs has provided Product #103B (Cholera Toxin B Subunit (Choleragenoid) from Vibrio cholerae) and Product #104 (Cholera Toxin B Subunit (Choleragenoid) from Vibrio cholerae in Low Salt).  The former has been discontinued.  The latter may be substituted for the former, if the researcher takes into account the difference in the buffer in which each product is provided and the amount of protein per vial.  When Product #103B was reconstituted with 200 μl of water, it contained 1 mg of protein in 0.05 M Tris, 0.2 M NaCl, 0.001 M Na2EDTA, 0.003 M NaN3, pH 7.5.  When Product #104 is reconstituted with 0.25 ml water, it contains 0.5 mg of Cholera Toxin B Subunit (choleragenoid) in 0.01 M Sodium Phosphate at pH 7.5.

PubMed ID: 19235216

Intranasal Delivery of Cholera Toxin Induces Th17-Dominated T-Cell Response to Bystander Antigens

Jee-Boong Lee, Ji-Eun Jang, Man Ki Song, Jun Chang
Product: Cholera Toxin (AZIDE-FREE) from Vibrio cholerae

Reagents:

CT and CTB were purchased from List Biological Laboratories (Campbell, CA). ...

Immunization:

For peptide immunization, 26106 OT-II splenocytes were i.v. injected into B6 mice, and then recipient mice were i.n. immunized with 40 mg of OVA peptide plus 30 mg of CpG ODN, 0.2 mg of CT, or 2 mg of CT or CTB in PBS. ...

Cytokine assays:

Dendritic cells (DCs) were generated from bone marrow of B6 or BALB/c mice by culturing in complete RPMI medium containing 10% FBS supplemented with 10 ng/ml recombinant GM-CSF and IL-4 (R&D, Minneapolis, MN). After 7 days of culture, non-adherent cells were harvested by gentle pipetting, and DCs were enriched by density gradient centrifugation over Percoll medium. After purification, DCs were further incubated with PBS or CT (100 ng/ml) for 3 days and the culture supernatants were harvested at the indicated time points. ...

 

Author did not specify which Cholera toxin was utilized.  List Labs Product #101B/C - Cholera Toxin from Vibrio Cholerae has been discontinued; however, List Labs still provides Product #100B - Cholera Toxin (AZIDE-FREE) from Vibrio cholerae.  

To follow, are our recommendations for transitioning from Product #101B/C to Product #100B:

Product #101B, Cholera Toxin from Vibrio cholerae (1 mg), has been discontinued.  Product #100B, Cholera Toxin (AZIDE-FREE) from Vibrio cholerae, may be substituted for Product #101B, if the user accounts for the formulation differences.  Both products contain the same quantity of toxin per vial (1.0 mg).  For Product #100B, reconstitution with 1.0 mL of water results in a 1 mg/mL toxin solution in a buffer with the following component concentrations:  0.05 M Tris, 0.2 M NaCl, 0.001 M Na2EDTA at pH 7.5.  For Product #101B, reconstitution with half that amount of water (0.5 mL) resulted in a 2 mg/mL toxin solution with the same concentrations of the buffer components, and in addition, Product #101B contained 0.003 M NaN3.  The user may add the sodium azide preservative to product #100B, if desired.

Product #101C, Cholera Toxin from Vibrio cholerae (2 mg), has been discontinued.  Product #100B, Cholera Toxin (AZIDE-FREE) from Vibrio cholerae, may be substituted for #101C, if the researcher accounts for the formulation differences.  While Product #101C contained 2.0 mg per vial, Product #100 contains 1.0 mg per vial.  When Product #100B is reconstituted with 1.0 mL of water, the resulting 1 mg/mL toxin solution will have buffer components with the following concentrations:  0.05 M Tris, 0.2 M NaCl, 0.001 M Na2EDTA at pH 7.5.  For Product #101C, reconstitution with less than half that amount of water (0.4 mL) resulted in a 5 mg/mL toxin solution in the same buffer, and in addition, Product #101C contained 0.003 M NaN3.  The user may add the sodium azide preservative to product #100B, if desired. 

Large quantities of Product #101B or Product #101C may be obtained through Bulk Requests and Customer Orders

 

Author did not specify which CTB was utilized.  List Labs has provided Product #103B (Cholera Toxin B Subunit (Choleragenoid) from Vibrio cholerae) and Product #104 (Cholera Toxin B Subunit (Choleragenoid) from Vibrio cholerae in Low Salt).  The former has been discontinued.  The latter may be substituted for the former, if the researcher takes into account the difference in the buffer in which each product is provided and the amount of protein per vial.  When Product #103B was reconstituted with 200 μl of water, it contained 1 mg of protein in 0.05 M Tris, 0.2 M NaCl, 0.001 M Na2EDTA, 0.003 M NaN3, pH 7.5.  When Product #104 is reconstituted with 0.25 ml water, it contains 0.5 mg of Cholera Toxin B Subunit (choleragenoid) in 0.01 M Sodium Phosphate at pH 7.5.

PubMed ID: 19360100