Dotmatics

Novalix ASMS Kd Demo

The Novalix ASMS Kd Demo details Virscidian's standalone manual data workflow involving data acquisition, transfer, and analysis with Analytical Studio-Pro, specifying required and optional metadata formats for sample and assay information, alongside SMS+Express services that provide automated data management, analysis, and storage with APIs and sample list building for affinity selection mass spectrometry experiments.

Virscidian Environments

Standalone Manual Data Workflow

  • Data acquisition
  • Data transfer
  • Data analysis by Analytical Studio-Pro
  • Data output

Metadata Requirements (Standalone Environments)

  • Required Metadata:

    • Workflow name: WN! (e.g., WN!KD-std)
    • Assay type: AT! (e.g., AT!ASMS)
    • Plate layout: PL! (e.g., PL!01TL,XY,NL,1+16,2+24)
    • Sample location: L! (e.g., L!C2)
    • Protein name: PROTNAME! (e.g., PROTNAME!Enzyme1)
    • Protein concentration: PROTCONC! (e.g., PROTCONC!10)
    • Protein concentration units: PROTCONCUNITS! (e.g., PROTCONCUNITS!uM)
    • Note: Compound concentration metadata is stored in the targets file.

  • Nice to Have Metadata:

    • Reaction time: RXNT! (e.g., RXNT!1)
    • Reaction time units: RXNTUNITS! (e.g., RXNTUNITS!hour)
    • Replicate: REP! (e.g., REP!1)

  • Example SI String (for comments section of injection sequence):

    SI=WN!KD-STD:PL!01TL,XY,NL,1+16,2+24:L!B04:SAM!MSOnly:SCM!MSOnly:SUBCONC!0.041:SUBCONCUNITS!uM:SN!MSC2036175B-6:PROTNAME!Protein1:F!C22H24N6O4S:

Example of Sample Info Metadata Format

  • SI=AT!ASMS:L!A09:PL!01TL,XY,NL,1+8,2+12:BAR!ABC123
    • SI denotes the start of a sample info string, followed by key-value pairs.
      1. 1.AT: assay type (e.g., ASMS for affinity selection MS)
      2. 2.L: well location (e.g., A09)
      3. 3.PL: plate layout (e.g., 01TL,XY,NL,1+8,2+12)
      4. 4.BAR: barcode for tracking (e.g., ABC123)

SMS + Express Sample Management Services

  • Analytical Studio-Express
    • Metadata, sample info, results management
    • Data analysis & processing
    • APIs available
    • Data storage on an MS SQL database server
    • Automated data transfer by Instrument Agent
    • Sample List Builder creates sample lists / work lists
    • Data output acquisition
    • Analytical Studio-Pro (injection sequences)

Metadata (for SMS+Express Environments)

  • Fields:

    • ASMS campaign number
    • Plate Barcode
    • PLATE_TYPE
    • Well
    • Compound ID
    • Formula
    • Adduct Override
    • Batch
    • IS Flag
    • Mixture
    • RT (min)
    • RT Window (min)
    • Tracking ID
    • Concentration
    • Concentration Units
    • Protein name
    • Protein Concentration
    • Protein Concentration Units

  • Example Data:

    • Kd Demo 001, 384, B01, Cpd-001, C22H24N6O4S, 0, uM
    • Kd Demo 001, 384, B04, Cpd-001, C22H24N6O4S, 0.041, uM
    • Kd Demo 001, 384, B05, Cpd-001, C22H24N6O4S, 0.123, uM
    • Kd Demo 001, 384, B06, Cpd-001, C22H24N6O4S, 0.37, uM

  • Metadata is mapped from CSV to SMS database.

  • Metadata is referenced by SMS and Express Databases.

  • Automated data transfer by Instrument Agent.

  • Sample List Builder creates sample lists / work lists.

  • Metadata written by SMS.

Kd Analysis

Experiment Setup

  • Plate layouts are shown for multiple compounds and concentrations.
  • Example: Compounds 1–5 distributed across plate rows and columns.

Kd Modeling

  • MS response is measured as peak area or peak intensity.

  • Bmax: Maximal MS response where all binding sites are occupied.

  • Kd: Compound concentration at half of Bmax.

  • Model fit equation:

    y = (Bmax * x) / (Kd + x)

    • y: MS response
    • x: Compound concentration

Data Suitability: Curve Fitting

  • Curve fitting metrics:

    • MS readout
    • Bmax
    • Kd
    • R² (how well data fits the model)

  • Fit equation:

    y = (Bmax * x) / (Kd + x)

Data Suitability: Linear Fitting

  • Linear fitting metrics:

    • MS readout
    • Bmax
    • R² (fit to a line)
    • Slope
    • Y-Intercept
    • Intercept to Bmax ratio

  • Linear fit equation:

    y = (Bmax * x) / (Kd + x)

Intercept to Bmax Ratios

  • Values approaching 1: Not enough low concentration data points.
  • Values approaching 0 (or negative): Not enough high concentration data points or a non-specific Kd binder.

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