🔬 Pipette Calibration (Volumetric)

Metrology PN-EN ISO 8655

Overview

STATUS (as of 06.11.2026). PN-EN ISO 8655-6:2022-11 (English version, published on 08.11.2022) implements EN ISO 8655-6:2022 and ISO 8655-6:2022 and replaced PN-EN ISO 8655-6:2003 with the corrigendum AC:2009. ISO 8655-6:2022 is the second edition (April 2022, corrected version June 2022), which replaced ISO 8655-6:2002 with the corrigendum Cor 1:2008. Changes: new uncertainty requirements for the test equipment (Tables 1 and 2, in conjunction with ISO/TR 20461), a new Clause 4 “General requirements”, a new Formula (2) based on ISO 4787, Annex B deleted. The whole ISO 8655 series (parts 1–9; in PKN editions 2022-11; part 7 has the amendment A1:2025-06) was issued in the same year. The maximum permissible errors of pipettes are given in PN-EN ISO 8655-2:2022-11 (English version, published on 07.11.2022, replaced PN-EN ISO 8655-2:2003; in ISO 8655-2:2022 Tables 1 and 2 were revised and Table 3 added).

SCOPE (ISO 8655-6, clause 1). Gravimetric reference procedure for the determination of volume of piston-operated volumetric apparatus (pipettes, burettes, dilutors, dispensers, syringes) — complete systems including tips and other parts involved in the measurement, delivered (Ex) or contained (In). Whoever claims conformity with the standard shall meet all its requirements (clause 4).

CONDITIONS (clauses 5–7). Distilled or deionised water of at least grade 3 according to ISO 3696, at a temperature within ±0.5 °C of the air temperature. Draught-free room: relative humidity 45–80 %, temperature (20 ± 3) °C with a maximum variation of ±0.5 °C during the test; the conditions shall be maintained for at least 2 h before the test, and the pipette, equipment and water shall reach equilibrium. The balance is chosen according to the nominal volume from Table 1 (e.g. 20–200 µl: resolution 0.01 mg, repeatability 0.025 mg, expanded uncertainty in use 0.05 mg; 200 µl–10 ml: 0.1 / 0.2 / 0.4 mg) so that the uncertainty of the balance is less than one quarter of the maximum permissible systematic error. Thermometer for liquids: resolution 0.1 °C, uncertainty 0.2 °C; thermometer for air 0.1 / 0.3 °C; hygrometer 1 / 5 %; barometer 0.1 / 1 kPa. Evaporation at small volumes (< 50 µl) is determined experimentally and corrected; the test cycle time shall be short and regular.

PROCEDURE (clause 8). Fixed-volume pipette — the nominal volume is tested; variable-volume pipette — at least the nominal volume, 50 % of nominal and the lower limit of the range or 10 % of nominal (whichever is greater). At least 10 measurements for each volume. At the beginning and at the end the water temperature is recorded, as well as the air temperature and pressure and the humidity. Air-displacement pipettes: always forward pipetting; after fitting the tip, the test liquid is aspirated five times to reach humidity equilibrium in the pipette; the set volume is not changed during the series.

CALCULATIONS (clause 9). Evaporation correction, conversion of the corrected weighing values to volume — by the general formula or the Z factor (from water temperature and air pressure, examples in informative Annex A), mean volume, systematic error, random error and measurement uncertainty. The report includes, among others, the ambient conditions, including humidity.

WHAT WE DO NOT GIVE HERE. The available text of ISO 8655-6 ends at the beginning of the test cycle (8.3.2 d); further steps, the formulas of clause 9 and the tables of maximum permissible errors of ISO 8655-2:2022 (Tables 1–3, revised compared with the 2002 edition) are outside it — we therefore give no permissible systematic and random errors and no tip immersion depth (Table 3 of part 6).

Method principle

Water is dispensed with the pipette into a weighing vessel on a balance whose parameters suit the nominal volume, and each portion is weighed. The weighing values corrected for evaporation are converted to volume by the general formula or the Z factor, which takes account of the water temperature and air pressure. From at least 10 measurements for each volume the mean volume, the systematic and random errors and the uncertainty are calculated, and the result is compared with the maximum permissible errors of ISO 8655-2.

Applications

• Calibration and testing of single- and multi-channel air-displacement (type A) and positive-displacement (type D) piston pipettes according to ISO 8655-2
• Determination of the volume of burettes, dilutors, dispensers and manually operated precision syringes (ISO 8655-3, -4, -5, -9) by the same gravimetric procedure
• Check of a pipette after adjustment — subsequent adjustment requires reference to the procedure of ISO 8655-6, ISO 8655-7 or ISO 8655-8 (ISO 8655-2, 5.3)
• Tests by manufacturers, calibration laboratories, test houses and users (introduction to the ISO 8655 series)

Key parameters

Standard

Standard number

PN-EN ISO 8655-6:2022-11 / PN-EN ISO 8655-2:2022-11

Title (PL)

Tłokowe przyrządy do pomiaru objętości — Część 6: Referencyjna procedura pomiaru grawimetrycznego do wyznaczania objętości / Część 2: Pipety

Title (EN)

Piston-operated volumetric apparatus — Part 6: Gravimetric reference measurement procedure for the determination of volume / Part 2: Pipettes

Step-by-step procedure

1. Equilibration

   The pipette, equipment, exchangeable parts and water reach thermal equilibrium; room conditions (20 ± 3) °C and 45–80 % humidity for at least 2 h before the test.

   ⏱ Time: ≥ 2 h • 🌡 Temperature: (20 ± 3) °C

2. Volume setting

   Variable-volume pipette — set the test volume; do not alter it during all the measurements of the series.

3. Weighing vessel

   Clean vessel with water covering the bottom, on a balance selected according to Table 1.

4. Recording conditions

   At the start and at the end of the measurements — water temperature; in addition air temperature, barometric pressure and relative humidity.

5. Tip and humidity equilibrium

   Fit the selected tip and aspirate the water five times to reach humidity equilibrium of the air in the pipette; always forward pipetting.

6. Measurements

   At least 10 deliveries into the weighing vessel with the weighing values recorded; the cycle time short and regular (evaporation correction). Hand warmth transferred to the pipette causes a systematic drift in the series (the delivered volume decreases — ISO 8655-2, 6.2). Further steps of the cycle (8.3.2 e onwards) — according to the text of the standard.

7. Further volumes

   For a variable-volume pipette — also 50 % of nominal and the lower limit of the range or 10 % of nominal.

8. Calculations

   Evaporation correction, conversion to volume (general formula or Z factor), mean volume, systematic error, random error, measurement uncertainty; comparison with the maximum permissible errors of ISO 8655-2.

9. Report

   Results, uncertainty and ambient conditions, including relative humidity; if delivery followed the manufacturer’s instructions instead of the procedure of the standard — a description of that procedure.

Required equipment and apparatus

Reagents, media and consumables

Health and safety (OHS)

• Tests according to the ISO 8655 series are intended to be carried out by trained personnel (introduction to ISO 8655-6)

Frequently asked questions