Method Development Workflow
Creating an acquisition method in MassHunter involves defining which elements to measure, in which cell gas mode, and with what acquisition parameters. The Agilent 8900's Method Wizard simplifies this process significantly.
Step 1 — Element Selection
Navigate to the Acquisition tab in MassHunter. Select the elements you wish to analyse. For pharmaceutical heavy metal testing per USP ⟨232⟩/⟨233⟩, the minimum required elements are:
| Element | Symbol | Mass (m/z) | Recommended Mode | Note |
|---|---|---|---|---|
| Arsenic | As | 75 → 91 | O₂ MS/MS (mass-shift) | Measured as AsO⁺ to avoid ⁴⁰Ar³⁵Cl⁺ |
| Cadmium | Cd | 111, 114 | He mode or No Gas | Low interference at these masses |
| Lead | Pb | 208 | No Gas | Interference-free at high mass |
| Mercury | Hg | 202 | No Gas or He | Memory effect — requires extended rinse |
Step 2 — Cell Gas Mode Assignment
Assign the appropriate cell gas mode for each analyte. MassHunter's IntelliQuant Assistant can recommend optimal modes based on a preliminary scan, or you can manually assign modes based on known interferences.
Multi-Tune Acquisition
The Agilent 8900 supports multi-tune acquisition — the instrument can automatically switch between different cell gas modes during a single sample analysis. For example, within one acquisition:
- Tune 1 (No Gas): Pb, Hg, Bi (ISTD)
- Tune 2 (He Mode): Cd, Rh (ISTD)
- Tune 3 (O₂ MS/MS): As (→ AsO), Se (→ SeO), Ge (ISTD)
Cell gas switching is fast and automatic — there is no need to run separate methods for different modes.
Step 3 — Acquisition Parameters
Set the following key acquisition parameters for each element:
| Parameter | Typical Value | Purpose |
|---|---|---|
| Integration Time | 0.1 – 3.0 s per mass point | Longer = better precision, shorter = faster throughput |
| Replicates | 3 (standard) | Multiple measurements per sample for RSD calculation |
| Sweeps per Replicate | 100 (default) | Number of full mass scans averaged per replicate |
| Uptake / Stabilisation Time | 30–60 s | Time to aspirate sample before data collection begins |
| Rinse Time | 30–90 s | Time spent rinsing between samples (critical for Hg) |
Step 4 — Calibration Strategy
External Calibration with Internal Standard Correction
The standard approach for pharmaceutical ICP-MS analysis:
- Calibration standards: Prepare 5+ standards covering 0 – 10 ppb (or appropriate range) in the sample matrix (2% HNO₃). See Chapter 4 for details.
- Internal standard: Added online to all blanks, standards, and samples at constant concentration. ISTD signal is used to normalise analyte signals.
- Calibration curve: MassHunter plots ISTD-corrected signal vs. concentration. Linear regression (R² ≥ 0.999 preferred, ≥ 0.995 minimum).
- Verification: Run an independent CCV standard. Must recover within ±10%.
Step 5 — Batch / Sequence Setup
Navigate to the Sequence tab to create the sample run list. A typical analysis sequence follows this order:
| Position | Sample Type | Purpose |
|---|---|---|
| 1 | Calibration Blank | Establish zero baseline |
| 2–6 | Calibration Standards | Build calibration curve (ascending concentration) |
| 7 | CCV (mid-level std) | Verify calibration immediately after standards |
| 8 | CCB (blank) | Check for carryover |
| 9–18 | Samples | Unknown samples, spiked samples, duplicates |
| 19 | CCV | Mid-batch verification (every 10 samples) |
| 20 | CCB | Mid-batch blank check |
| 21–30 | Samples | Continue with remaining samples |
| 31 | CCV | End-of-batch verification |
| 32 | CCB | Final carryover check |
- Insert a CCV/CCB pair every 10 samples to monitor calibration stability.
- Place spiked samples alongside their unspiked counterparts for direct comparison.
- Use duplicate preparations to verify precision.
- Avoid using asterisks (*) in sample names — MassHunter may interpret them as wildcards.
- Assign autosampler rack positions carefully — double-check before starting the run.
Step 6 — Data Analysis
FullQuant Analysis
After acquisition, navigate to Data Analysis. MassHunter's FullQuant module automatically calculates sample concentrations from the calibration curve, applies internal standard corrections, and generates results tables.
Batch-at-a-Glance
The Batch-at-a-Glance interactive data table provides a real-time overview of:
- Sample concentrations (with pass/fail flagging against user-defined limits)
- Internal standard signal trends across the batch
- Calibration curve fit and individual point residuals
- Outlier flagging (built-in statistical checks)
- QC checks (CCV recovery, CCB levels)
Interpreting Results
| Check | Acceptance Criteria | Action if Fail |
|---|---|---|
| Calibration R² | ≥ 0.995 (ideally ≥ 0.999) | Re-prepare standards, check for contamination |
| CCV Recovery | 90–110% | Recalibrate if drift detected |
| CCB Level | ≤ LOQ | Investigate contamination, re-rinse |
| ISTD Stability | 70–130% of initial (in batch) | Investigate matrix suppression or drift |
| Sample RSD | ≤ 5% (3 replicates) | Re-run sample, check nebulizer |
| Spike Recovery | 70–150% (per USP ⟨233⟩) | Investigate matrix, re-prepare if needed |
Reporting
MassHunter can generate automated reports in customisable formats. Ensure reports include:
- Calibration curve parameters (slope, intercept, R²)
- Sample results (concentration, RSD, ISTD recovery)
- QC summary (CCV, CCB, spike recovery)
- Instrument tune/performance data
- Method parameters (elements, modes, integration times)