Sole_Source_Justification_as_required_by_FAR_13_501a_Redacted_2.pdf

SOLE SOURCE JUSTIFICATION FOR ACQUISITION OVER THE

SIMPLIFIED ACQUISITION THRESHOLD

PR: 0040654087

1. Contracting Agency and Activity. The Department of the Interior, United States Geological Survey, plans to contract on a sole source basis. This document sets forth the justification and approval as required by FAR 13.501(a).

2. Nature of Action Being Approved.

The U.S. Geological Survey, Kansas Water Science Center’s Organic Geochemistry Research Laboratory (OGRL) has a requirement for a firm fixed price requirements contract with Gold Standard Diagnostics for 96-well plate enzyme-linked immunosorbent assay (ELISA) kits, cell lysis kits, and cyanotoxin standards.

3. Description of Supplies or Services.

The requirements and parts for each ELISA kit, cell lysing kits, and chemical standards are listed below. Several of the reagents are proprietary and every effort was made to identify what came in each kit. Listed part numbers are from Gold Standard Diagnostics.

Enzyme-linked immunosorbent assays (ELISA), chemical cell lysis kits, and cyanotoxin standards are described below. The cyanotoxins that are screened using ELISA include:

anatoxin-a, cylindrospermopsin, microcystin, and saxitoxins. The chemical cell lysis kits are used to lyse cells to release cyanotoxins contained within. The algal toxin standards, including dihydroanatoxin-a, homoanatoxin-a, 7-deoxy-cylindrospermopsin, 7-epi-cylindrospermopsin, 13C4-(+)-anatoxin-a; 15N5-cylindrospermopsin; 15N5-7-deoxy-cylindrospermopsin; and 15N5-7-epi-cylindrospermopsin are used for method development and to quantify cyanotoxins by high resolution mass spectrometry.

1. Anatoxin-a (VFDF) ELISA kit (p/n 520060):

a. Salinity: no matrix effects noted up to 37 PPT.

b. Methanol tolerance: <2.5% methanol

c. The results must be read using 450nm. The evaluation of the ELISA should be done using a 4-parameter curve fit with a 0.99 r2 as minimum acceptable curve fit.

d. The plates must be 96 wells and be readable using a SprectraMax ABS Absorbance Microplate Reader. The plate wells must be optically clear and the absorbance of the blank wells at 450nm should not vary more than 5% RSD across an entire plate.

e. Minimum Reporting Limit = 0.15μg/L; maximum reporting limit = 5.0μg/L.

f. Cross Reactivities: (+)-anatoxin-a – 100%; homoanatoxin-a – 124.8%; (-)-anatoxin-a 0.3%.

g. Required reagents and material: microtiter plate coated with secondary antibody;

lyophilized anatoxin-a-horseradish peroxidase enzyme conjugate; conjugate diluent;

lyophilized anti-anatoxin-antibody; antibody diluent; (+)-anatoxin-a calibration standards (six standards with a volume of 1ml each: 0, 0.15, 0.40, 1.0, 2.5, and 5.0μg/L); kit control standard (0.75μg/L); sample diluent (10x); wash buffer (5x); substrate (color) solution; and stop solution.

h. volume of sample required = 50μL.

i. Accuracy limits: kit control standard must be ≤ ±25% of the stated value. Precision limits:

the %RSD for all duplicates (blanks, controls, and samples) should be ≤ 35.4%.

2. Cylindrospermopsin ELISA kit (p/n 522011):

a. Salinity: must be less than 20%.

b. methanol tolerance: <20% methanol.

c. The following substances can have no significant effect on assay results: calcium chloride, magnesium chloride, magnesium sulfate, manganese sulfate, and sodium chloride up to 10,000 ppm; calcium sulfate, ferric sulfate, potassium phosphate, sodium nitrate, and zinc sulfate up to 1,000 ppm; copper chloride up to 100 ppm; Lugol’s solution up to 0.01%.

Additionally, the presence of sodium thiosulfate (up to 0.5mg/ml) and ascorbic acid (up to 1mg/ml) should not affect the assay.

d. The results must be read using 450nm. The evaluation of the ELISA should be done using a 4-parameter curve fit with a 0.99 r2 as minimum acceptable curve fit.

e. The plates must be 96 wells and be readable using a SpectraMax ABS Absorbance Microplate Reader. The plate wells must be optically clear and the absorbance of the blank wells at 450nm should not vary more than 5% RSD across an entire plate.

f. minimum reporting limit = 0.05μg/L; maximum reporting limit = 2.0μg/L.

g. cross reactivities: cylindrospermopsin – 100%; deoxy-cylindrospermopsin – 112%; and 7-epi-cylindrospermopsin – 157%.

h. Required reagents: microtiter 96 well plate; calibration standards (1.5ml each, seven in total, 0, 0.05, 0.10, 0.25, 0.50, 1.0, and 2.0μg/L); kit control (1.5ml, 0.75μg/L); sample diluent; lyophilized cylindrospermopsin-Horse Radish Peroxidase enzyme conjugate;

conjugate diluent; Empty amber HDPE bottle for combining reconstituted Enzyme Conjugate; Antibody Solution (rabbit anti-Cylindrospermopsin); Wash Buffer (5X) Concentrate; Substrate (Color) Solution (TMB); and stop solution.

i. volume of sample required = 50μL

j. Accuracy limits: kit control standard must be ≤ ±25% of the stated value. Precision limits:

the %RSD for all duplicates (blanks, controls, and samples) should be ≤ 35.4%.

3. Microcystin/Nodularins (ADDA) SAES ELISA kit (p/n 520011SAES):

a. Salinity: no matrix effects observed up to 38 PPT salinity.

b. methanol tolerance: samples must not contain methanol concentrations > 1%

c. No matrix effects observed with samples which have been treated with sodium thiosulfate (≤ 10 mg/mL) or ascorbic acid (≤ 1 mg/mL).

d. The results must be read using 450nm. The evaluation of the ELISA should be done using a 4-parameter curve fit with a 0.99 r2 as minimum acceptable curve fit.

e. The plates must be 96 wells and be readable using a SpectraMax ABS Absorbance Microplate Reader. The plate wells must be optically clear and the absorbance of the blank wells at 450nm should not vary more than 5% RSD across an entire plate.

f. minimum reporting limit = 0.10μg/L; maximum reporting limit = 5.0μg/L.

g. the assay must exhibit approximately equimolar cross reactivity (e.g. ~100%) for all cyanobacterial cyclic peptides toxic congeners including: microcystin-LR, microcystin-YR, microcystin-LF, desmethyl-microcystin-RR, nodularin-R, microcystin-RR, microcystin- LW, microcystin-LA and desmethyl-microcystin-LR.

h. Required reagents: microtiter 96 well plate coated with analog of microcystins conjugated to a protein; calibration standard (1.5ml each, six in total: 0, 0.05, 0.15, 0.4, 1.5, 5.0μg/L);

kit control (1.5ml, 0.75μg/L); Sample Diluent; Microcystins-ADDA SAES Antibody Solution; Microcystins-ADDA SAES Conjugate Solution; Wash Buffer (5X) Concentrate;

Substrate (Color) Solution (TMB); and stop solution.

i. volume of sample required = 50μL

j. Accuracy limits: kit control standard must be ≤ ±25% of the stated value. Precision limits:

the %RSD for all duplicates (blanks, controls, and samples) should be ≤ 35.4%.

4. Saxitoxin/Lyngbyatoxin (PSP) ELISA kit (p/n 52255B):

a. Salinity: no matrix effects observed up to 27 PPT

b. methanol tolerance: must be less than 20%

c. The results must be read using 450nm. The evaluation of the ELISA should be done using a 4-parameter curve fit with a 0.99 r2 as minimum acceptable curve fit.

d. The plates must be 96 wells and be readable using a SpectraMax ABS Absorbance Microplate Reader. The plate wells must be optically clear and the absorbance of the blank wells at 450nm should not vary more than 5% RSD across an entire plate.

e. minimum reporting limit = 0.02μg/L; maximum reporting limit = 0.40μg/L.

f. cross reactivity – saxitoxin -100%; Decarbamoyl STX - 29%; GTX 2 & 3 - 23%; GTX-5B

- 23%; Lyngbyatoxin - 13%; Sulfo GTX 1 & 2 - 2.0%; Decarbamoyl GTX 2 & 3 - 1.4%;

Neosaxitoxin - 1.3%; Decarbamoyl Neo STX - 0.6%; and GTX 1 & 4 <0.2%.

g. Required reagents: Microtiter plate coated with a second antibody (anti-rabbit); calibration standards (6): 0, 0.02, 0.05, 0.1, 0.2, 0.4 ng/mL STX-diHCl, 1.5 mL each; kit control 0.075ng/mL STX-diHCl, 1.5 mL; Antibody Solution (rabbit anti-Saxitoxin); Saxitoxin- Horse Radish Peroxidase Conjugate Solution; Sample Diluent (10X) Concentrate; wash buffer (5X); substrate color solution (TMB); and stop solution.

h. volume of sample required = 50μL

i. Accuracy limits: kit control standard must be ≤ ±25% of the stated value. Precision limits:

the %RSD for all duplicates (blanks, controls, and samples) should be ≤ 35.4%.

5. QuickLyse Cell lysis kit (p/n 529911):

The chemical cell lysis kits are used to lyse cells to release cyanotoxins contained within cells. The AET laboratory is using an earlier NOAA validated method and therefore using the same lysing kits is vital for data comparison. The ingredients for this kit are proprietary and therefore cannot be listed here.

6. Cyanotoxin standards and isotopically labeled standards minimum/maximum reporting levels, calibration standard concentrations, check standards concentrations, and sample volume requirements then Gold Standard Diagnostic ELISA kits.

The Gold Standard Diagnostic ELISA kits we have been using also have had a consistent accuracy (≤ ±25% of the stated concentration of the control standards) and precision (%RSD for duplicates ≤ 35.4%) values that may differ from other manufactures. The kits’ minimum reporting level, acceptable R2 value of calibration curve, and calibration range is shown in the table below.

These changes make comparing historical data from a specific site or project difficult and will complicate long term studies. Different manufacturers have different tolerances for salinity, methanol, and other possible interfering matrix chemicals. For the kits produced by Gold Standard Diagnostics, the OGRL has validated these kits for sample preparation and matrix chemicals. Additionally, the ELISA kits currently in use by OGRL have been cross validated against mass spectrometry. If an alternative source for these kits is chosen, the validation experiments will have to be repeated which would cause either a delay of months before the kits would be put into use.

The OGRL AET lab group uses cyanotoxin standards to validate and optimize methods primarily on the high-resolution mass spectrometry, but they are occasionally used with other methods including ELISA. These standards include both unlabeled and isotopically labelled cyanotoxins.

The cyanotoxin standards purchased from Gold Standard Diagnostics include: dihydroanatoxin-a, homoanatoxin-a, 7-deoxy-cylindrospermopsin, 7-epi-cylindrospermopsin, 13C4-(+)-anatoxin-a;

15N5-cylindrospermopsin; 15N5-7-deoxy-cylindrospermopsin; and 15N5-7-epi-cylindrospermopsin.

As far as it is known, these cyanotoxin standards are only available from Gold Standard Diagnostics. The table below shows the minimum purity, isotope incorporation, and concentration or amount that have been available from Gold Standard Diagnostics.

Analyte Gold Standard Diagnostic Part

Number

Minimum Purity

Isotope Incorporation

Concentration or amount/vial

13C4-(+)-anatoxin-a 300505 98% >99% 10μg/ml dihydroanatoxin-a 300655 97% -- 10μg/ml homoanatoxin-a 301295 98% -- 4μg/ml 15N5-cylindrospermopsin 300504 99% >99% 10μg/ml 15N5-7-epi-cylindrospermopsin

300506 97% >99% 5μg

ELISA name Gold Standard

Diagnostic part number

Minimum reporting level

(μg/L) Calibration range (μg/L) Acceptable R2 value of calibration curve

Anatoxin-a (VFDF) ELISA kit 520060 0.15 0.15-5.0 0.99 Cylindrospermopsin ELISA kit 522011 0.05 0.05-2.0 0.99 Microcystin/Nodularins (ADDA) SAES ELISA kit 520011SAES 0.10 0.10-5.0 0.99 Saxitoxin/Lyngbyatoxin (PSP) ELISA kit 52255B 0.02 0.02-0.4 0.99