Quanterix_Statement_of_Work.pdf

SCOPE/STATEMENT OF WORK

Project Title: Calbindin D Assay Development

Tasks and Test Plan:

TASK A: Evaluate one established antibody pair for Calbindin D and determine preliminary lower limit of detection (LLOD).

TEST PLAN: Quanterix will develop prototype assay to Calbindin D protein, a biomarker target specified by NICHD. One antibody pair will be evaluated using the HD-1/X Analyzer.

Standard reagent preparation and antibody screening procedures will be conducted. The preliminary assay lower limit of detection (LLOD) will be determined using recombinant protein or peptide. To facilitate screening an abbreviated calibration curve may be used.

Quanterix will purchase 600 µg each of the capture and detection antibodies and 10 µg of Calbindin D recombinant protein or peptide. Quanterix will provide the homebrew kits, required consumables and all other reagents needed to run the HD-1/X analyzer and develop an assay.

Milestones: Task A Milestone: A PowerPoint slide deck detailing assay evaluations using antibodies tested. The best performing antibody capture-reporter combinations will be identified which meets proof of-concept requirements.

TASK B: Following the determination of the preliminary LLOD using the established antibody pair, continue assay optimization and matrix evaluation on SR-X instrument.

TEST PLAN: The established antibody pair will be further evaluated using the SR-X instrument. Optimization may include additional exploration of antibody loading on beads, assessment of conjugation chemistry, stoichiometry and reaction buffers. In addition, optimization of antibody biotinylation level as well as detector antibody concentration may be evaluated. Detector diluents and assay wash conditions (antibody pair dependent) will also be assessed to minimize non-specific background (NSB).

Milestones: Task B Milestone: Completion of assay optimization and matrix evaluation, and report detailing results.

TASK C: Using the optimal assay conditions determined in Task B, perform validation studies to further define assay performance on SR-X instrument.

TEST PLAN: The purpose of validating prototype Simoa assay is to characterize assay performance and demonstrate its suitability to quantify the levels of target protein in human sample. A scale-up of reagents and crossover study using optimal conditions identified in Task B will be required. Additional antibody will be required to proceed with validation efforts. Quanterix will purchase 1 mg of capture antibody and 0.6 mg of detection antibody as well as sufficient CSF samples to complete experiments defined in Tasks C and D.

Quanterix will provide the homebrew kits, required consumables and all other reagents needed.

Milestones: Task C Milestone: Completion of validation studies and report detailing these results.

TASK D: Production of assay materials sufficient for the analysis of 300 human CSF samples Quanterix will utilize a homebrew development kit to prepare materials sufficient for characterization and subsequent conversion to minimum of ten individual calbindin D kits. Quanterix requests the same lot of antibody used for previous development work to minimize additional variability.

Calbindin D kits and all other supplemental information needed to run the SR-X will be provided to NICHD for sample testing.

Task D Milestone: Completion of calbindin D assay kit production and report detailing these results additional variability.

Summary Tasks:

TASK A: Evaluation of an established antibody pair for calbindin D protein, a biomarker specified by NICHD. Recombinant protein will be used to establish calibration curves and determine assay sensitivity lower limit of detection (LLOD).

TASK B: Following the determination of the preliminary LLOD using the established antibody pair (Task A), continue with assay development and optimization. Preliminary spike recovery, dilutional linearity and lower limit of quantification (LLOQ) determination studies will be conducted on SR-X to assess performance in human CSF.

TASK C: Using the optimized assay conditions identified in Task B, fit-for-purpose validation studies will be conducted to qualify the assay performance on SR-X, including sensitivity, linearity, recovery, dynamic range, and precision.

TASK D: Production of assay materials sufficient for the analysis of 300 human CSF samples (estimate ten 96-well plates). Materials will be prepared in kit format and provided to NICHD.

Project Timelines:

Task A will be completed within 3 weeks from the time all the materials are received.

Task B will start upon successful completion of Task A and will be completed within 4 weeks from the time all materials are received.

Task C will start upon successful completion of Task B and will be completed within 4-6 weeks from the time all materials are received.

Task D will start upon successful completion of Task C and will be completed within 3 weeks from the time all materials are received.

Project Deliverables:

Task A-D:

• Relevant test protocols

• Relevant data generated during testing

• Description of Data analysis methods / statistics, including, but not limited to, interpretation of the Data when the Data is “technology / device” specific.