steps for sample preparation, data recording, regression analysis, and documentation to demonstrate the method’s ability to produce results that are directly proportional to the concentration of analyte across a specified range.

2. Scope

This procedure is applicable to all analytical methods under development within the Analytical Method Development (AMD) department for assay, related substances, dissolution, and content uniformity methods. It is applicable for both chromatographic and non-chromatographic techniques.

3. Responsibilities

• Analyst: Prepares solutions, performs injections, and records responses.
• Data Reviewer: Reviews linearity data, verifies calculations, and confirms compliance with acceptance criteria.
• QA Personnel: Ensures the documented data table is audit-ready and conforms with ICH and GMP requirements.

4. Accountability

The Head of Analytical Method Development is accountable for ensuring the accuracy, completeness, and compliance of linearity evaluations and associated data tables as part of method validation reports.

5. Procedure

5.1 Preparation of Linearity Solutions

1. Select a concentration range typically from 50% to 150% of the target concentration. Include a minimum of 5 concentration levels (e.g., 50%, 75%, 100%, 125%, 150%).
2. Prepare each concentration level using a validated dilution technique and record dilution factors.
3. Label and store the linearity samples appropriately as per stability data.

5.2 Data Collection

1. Inject each level (preferably in triplicate) into the chromatographic system or record absorbance/response.
2. Ensure system suitability parameters are met before proceeding with linearity injections.
3. Record retention time (if applicable), area, and response for each injection in a raw data sheet.

5.3 Preparation of Linearity Data Table

1. Create a structured table with the following columns:
   • Concentration (% of target)
   • Theoretical Concentration (μg/mL)
   • Peak Area or Response
   • Average Response
   • Standard Deviation
   • Relative Standard Deviation (RSD%)
2. Calculate average area and % RSD for each concentration level.
3. Plot Concentration vs. Average Area or Response to generate the calibration curve.

5.4 Regression Analysis

1. Perform linear regression using suitable software (e.g., Microsoft Excel, Empower, or validated LIMS software).
2. Calculate:
   • Slope (m)
   • Intercept (c)
   • Coefficient of Determination (R²)
3. Acceptable criteria:
   • R² ≥ 0.998 for assay methods
   • R² ≥ 0.990 for impurity methods

5.5 Documentation and Evaluation

1. Attach printouts of chromatograms or spectrophotograms for each level.
2. Include the regression equation and linearity graph in the method validation report.
3. Document any observations related to carryover, peak distortion, or baseline drift.

6. Abbreviations

• RSD: Relative Standard Deviation
• QA: Quality Assurance
• ICH: International Council for Harmonisation
• LOD: Limit of Detection
• LOQ: Limit of Quantitation

7. Documents

1. Linearity Calculation Sheet – Annexure-1
2. Chromatograms/Spectra Records – Annexure-2
3. Regression Analysis Summary – Annexure-3

8. References

• ICH Q2(R1) – Validation of Analytical Procedures: Text and Methodology
• FDA Guidance for Industry – Analytical Procedures and Methods Validation
• 21 CFR Part 211 – Current Good Manufacturing Practice

9. SOP Version

Version: 2.0

10. Approval Section

	Prepared By	Checked By	Approved By
Signature
Date
Name	Swati Mehra	Karan Vyas	Dr. Harshita Goyal
Designation	Analyst – AMD	QA Reviewer	Head – AMD
Department	Analytical Method Development	Quality Assurance	Analytical Method Development

11. Annexures

Annexure-1: Linearity Calculation Sheet

Conc. (%)	Theoretical Conc. (μg/mL)	Avg. Area	SD	RSD (%)
50	10	10235	45	0.44
75	15	15410	60	0.39
100	20	20580	80	0.39
125	25	25740	92	0.36
150	30	30790	110	0.36

Annexure-2: Chromatograms/Spectra

Attach all raw data printouts or electronic files from HPLC or UV systems used in linearity evaluation.

Annexure-3: Regression Analysis Summary

Linearity Equation: y = 1023.6x + 50.2
R² = 0.9991
Slope and intercept within expected range based on method requirements.

Revision History:

Revision Date	Revision No.	Details	Reason	Approved By
01/06/2025	2.0	Expanded linearity procedure and included annexure formats	Annual SOP Review	Dr. Harshita Goyal
15/03/2022	1.0	Initial version	New SOP	QA Head