EN ISO 23783-2:2023
(Main)Automated liquid handling systems - Part 2: Measurement procedures for the determination of volumetric performance (ISO 23783-2:2022)
Automated liquid handling systems - Part 2: Measurement procedures for the determination of volumetric performance (ISO 23783-2:2022)
This document specifies procedures for the determination of volumetric performance of automated liquid handling systems (ALHS), including traceability and estimations of measurement uncertainty of measurement results.
This document is applicable to all ALHS with complete, installed liquid handling devices, including tips and other essential parts needed for delivering a specified volume, which perform liquid handling tasks without human intervention into labware.
NOTE For terminology and general requirements of automated liquid handling systems, see ISO 23783-1. Determination, specification, and reporting of volumetric performance of automated liquid handling systems is described in ISO 23783-3.
Automatisierte Flüssigkeitsdosiersysteme - Teil 2: Messverfahren zur Bestimmung der volumetrischen Leistung (ISO 23783-2:2022)
Dieses Dokument legt Verfahren zur Bestimmung der volumetrischen Leistung von automatisierten Flüssigkeitsdosiersystemen (ALHS), einschließlich Rückverfolgbarkeit und Abschätzung der Messunsicherheit von Messergebnissen fest.
Dieses Dokument ist anwendbar für alle ALHS mit vollständigen, installierten Flüssigkeitsdosiersystemen, einschließlich Spitzen und anderer wesentlicher Teile, die für die Dosierung eines bestimmten Volumens benötigt werden und die Flüssigkeitsdosier-Aufgaben ohne menschliches Zutun in Laborgeräten durchführen.
ANMERKUNG Zur Terminologie und den allgemeinen Anforderungen an automatisierte Flüssigkeitsdosiersysteme siehe ISO 23783 1. Die Bestimmung, Spezifikation und Dokumentation der volumetrischen Leistung von automatisierten Flüssigkeitsdosiersystemen wird in ISO 23783 3 beschrieben.
Systèmes automatisés de manipulation de liquides - Partie 2: Procédures de mesure pour la détermination des performances volumétriques (ISO 23783-2:2022)
Le présent document spécifie les procédures pour la détermination des performances volumétriques des systèmes automatisés de manipulation de liquides (ALHS), y compris la traçabilité et les estimations de l’incertitude de mesure des résultats obtenus.
Le présent document s’applique à tous les ALHS dans lesquels sont installés des dispositifs complets de manipulation de liquides, y compris les cônes et autres composants essentiels nécessaires à la distribution d’un volume spécifié, qui exécutent des tâches de manipulation de liquides avec du matériel de laboratoire, sans intervention humaine.
NOTE Pour la terminologie et les exigences générales applicables aux systèmes automatisés de manipulation de liquides, voir l’ISO 23783-1. La détermination, la spécification et le compte-rendu des performances volumétriques des systèmes automatisés de manipulation de liquides sont décrits dans l’ISO 23783-3.
Avtomatizirani sistemi za ravnanje s tekočinami - 2. del: Merilni postopki za določanje prostorninske zmogljivosti (ISO 23783-2:2022)
Ta dokument določa postopke za določanje volumetrične zmogljivosti avtomatiziranih sistemov za ravnanje s tekočinami (ALHS), vključno s sledljivostjo in ocenami merilne negotovosti merilnih rezultatov.
Ta dokument se uporablja za vse avtomatizirane sisteme za ravnanje s tekočinami s popolnimi, nameščenimi napravami za ravnanje s tekočinami, vključno s konicami in drugimi bistvenimi deli, ki so potrebni za dovajanje določene količine. Gre za sisteme, ki izvajajo opravila ravnanja s tekočinami brez človeškega posredovanja v laboratorijsko opremo.
OPOMBA: Za terminologijo in splošne zahteve avtomatiziranih sistemov za ravnanje s tekočinami glej standard ISO 23783-1. Določanje, specifikacija in poročanje volumetrične zmogljivosti avtomatiziranih sistemov za ravnanje s tekočinami so opisani v standardu ISO 23783-3.
General Information
Standards Content (Sample)
SLOVENSKI STANDARD
01-december-2023
Avtomatizirani sistemi za ravnanje s tekočinami - 2. del: Merilni postopki za
določanje prostorninske zmogljivosti (ISO 23783-2:2022)
Automated liquid handling systems - Part 2: Measurement procedures for the
determination of volumetric performance (ISO 23783-2:2022)
Automatisierte Flüssigkeitsdosiersysteme - Teil 2: Messverfahren zur Bestimmung der
volumetrischen Leistung (ISO 23783-2:2022)
Systèmes automatisés de manipulation de liquides - Partie 2: Procédures de mesure
pour la détermination des performances volumétriques (ISO 23783-2:2022)
Ta slovenski standard je istoveten z: EN ISO 23783-2:2023
ICS:
17.060 Merjenje prostornine, mase, Measurement of volume,
gostote, viskoznosti mass, density, viscosity
71.040.20 Laboratorijska posoda in Laboratory ware and related
aparati apparatus
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
EN ISO 23783-2
EUROPEAN STANDARD
NORME EUROPÉENNE
October 2023
EUROPÄISCHE NORM
ICS 17.060; 71.040.20
English Version
Automated liquid handling systems - Part 2: Measurement
procedures for the determination of volumetric
performance (ISO 23783-2:2022)
Systèmes automatisés de manipulation de liquides - Automatisierte Flüssigkeitsdosiersysteme - Teil 2:
Partie 2: Procédures de mesure pour la détermination Messverfahren zur Bestimmung der volumetrischen
des performances volumétriques (ISO 23783-2:2022) Leistung (ISO 23783-2:2022)
This European Standard was approved by CEN on 25 September 2023.
CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this
European Standard the status of a national standard without any alteration. Up-to-date lists and bibliographical references
concerning such national standards may be obtained on application to the CEN-CENELEC Management Centre or to any CEN
member.
This European Standard exists in three official versions (English, French, German). A version in any other language made by
translation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC Management
Centre has the same status as the official versions.
CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia,
Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway,
Poland, Portugal, Republic of North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Türkiye and
United Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION
EUROPÄISCHES KOMITEE FÜR NORMUNG
CEN-CENELEC Management Centre: Rue de la Science 23, B-1040 Brussels
© 2023 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 23783-2:2023 E
worldwide for CEN national Members.
Contents Page
European foreword . 3
European foreword
The text of ISO 23783-2:2022 has been prepared by Technical Committee ISO/TC 48 "Laboratory
equipment” of the International Organization for Standardization (ISO) and has been taken over as
which is held by DIN.
This European Standard shall be given the status of a national standard, either by publication of an
identical text or by endorsement, at the latest by April 2024, and conflicting national standards shall be
withdrawn at the latest by April 2024.
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. CEN shall not be held responsible for identifying any or all such patent rights.
Any feedback and questions on this document should be directed to the users’ national standards body.
A complete listing of these bodies can be found on the CEN website.
According to the CEN-CENELEC Internal Regulations, the national standards organizations of the
following countries are bound to implement this European Standard: Austria, Belgium, Bulgaria,
Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland,
Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Republic of
North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Türkiye and the
United Kingdom.
Endorsement notice
The text of ISO 23783-2:2022 has been approved by CEN as EN ISO 23783-2:2023 without any
modification.
INTERNATIONAL ISO
STANDARD 23783-2
First edition
2022-08
Automated liquid handling systems —
Part 2:
Measurement procedures for
the determination of volumetric
performance
Systèmes automatisés de manipulation de liquides —
Partie 2: Procédures de mesure pour la détermination des
performances volumétriques
Reference number
ISO 23783-2:2022(E)
ISO 23783-2:2022(E)
© ISO 2022
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photocopying, or posting on
the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address below
or ISO’s member body in the country of the requester.
ISO copyright office
CP 401 • Ch. de Blandonnet 8
CH-1214 Vernier, Geneva
Phone: +41 22 749 01 11
Email: copyright@iso.org
Website: www.iso.org
Published in Switzerland
ii
ISO 23783-2:2022(E)
Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Abbreviated terms . 1
5 Measurement methods . 2
5.1 Overview of methods suitable for measuring ALHS performance . 2
5.2 Photometric methods . 9
5.2.1 Dual-dye ratiometric photometric method . 9
5.2.2 Single-dye photometric method . 9
5.2.3 Fluorescence method . 9
5.3 Gravimetric methods . 9
5.3.1 Single channel method . 9
5.3.2 Regression analysis . 10
5.4 Hybrid photometric/gravimetric method . 10
5.5 Dimensional methods . 10
5.5.1 Optical image analysis of droplets . 10
5.5.2 Optical image analysis of capillaries . 11
6 Equipment and preparation.11
6.1 Test equipment . 11
6.2 Manually operated single- and multi-channel pipettes .12
6.3 Preparation for testing .12
7 Thermal expansion .13
8 Traceability and measuring system uncertainty .13
8.1 Traceability . 13
8.2 Estimation of measuring system uncertainty . 13
8.2.1 Whole system approach . 13
8.2.2 Measurement model approach. 13
9 Reporting .14
Annex A (normative) Calculation of liquid volumes from balance readings .15
Annex B (normative) Dual-dye ratiometric photometric procedure .18
Annex C (normative) Single dye photometric procedure .24
Annex D (normative) Gravimetric procedure, single channel measurement .29
Annex E (normative) Gravimetric regression procedure .33
Annex F (normative) Photometric/gravimetric hybrid procedure .39
Annex G (normative) Optical image analysis of droplets .48
Annex H (normative) Fluorescence procedure.57
Annex I (normative) Optical image analysis of capillaries .70
Bibliography .76
iii
ISO 23783-2:2022(E)
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards
bodies (ISO member bodies). The work of preparing International Standards is normally carried out
through ISO technical committees. Each member body interested in a subject for which a technical
committee has been established has the right to be represented on that committee. International
organizations, governmental and non-governmental, in liaison with ISO, also take part in the work.
ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of
electrotechnical standardization.
The procedures used to develop this document and those intended for its further maintenance are
described in the ISO/IEC Directives, Part 1. In particular, the different approval criteria needed for the
different types of ISO documents should be noted. This document was drafted in accordance with the
editorial rules of the ISO/IEC Directives, Part 2 (see www.iso.org/directives).
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of
any patent rights identified during the development of the document will be in the Introduction and/or
on the ISO list of patent declarations received (see www.iso.org/patents).
Any trade name used in this document is information given for the convenience of users and does not
constitute an endorsement.
For an explanation of the voluntary nature of standards, the meaning of ISO specific terms and
expressions related to conformity assessment, as well as information about ISO's adherence to
the World Trade Organization (WTO) principles in the Technical Barriers to Trade (TBT), see
www.iso.org/iso/foreword.html.
This document was prepared by Technical Committee ISO/TC 48, Laboratory equipment.
This first edition of ISO 23783-2, together with ISO 23783-1 and ISO 23783-3, cancels and replaces
IWA 15:2015.
A list of all parts in the ISO 23783 series can be found on the ISO website.
Any feedback or questions on this document should be directed to the user’s national standards body. A
complete listing of these bodies can be found at www.iso.org/members.html.
iv
ISO 23783-2:2022(E)
Introduction
Globalization of laboratory operations requires standardized practices for operating automated
liquid handling systems (ALHS), communicating test protocols, as well as analysing and reporting
of performance parameters. IWA 15:2015 was developed to provide standardized terminology, test
protocols, and analytical methods for reporting test results. The concepts developed for, and described
in, IWA 15 form the foundation of the ISO 23783 series.
Specifically, this document addresses the needs of:
— users of ALHS, as a basis for calibration, verification, validation, optimization, and routine testing of
trueness and precision;
— manufacturers of ALHS, as a basis for quality control, communication of acceptance test specifications
and conditions, and issuance of manufacturer’s declarations (where appropriate);
— test houses and other bodies, as a basis for certification, calibration, and testing.
The tests established in this document should be carried out by trained personnel.
v
INTERNATIONAL STANDARD ISO 23783-2:2022(E)
Automated liquid handling systems —
Part 2:
Measurement procedures for the determination of
volumetric performance
1 Scope
This document specifies procedures for the determination of volumetric performance of automated
liquid handling systems (ALHS), including traceability and estimations of measurement uncertainty of
measurement results.
This document is applicable to all ALHS with complete, installed liquid handling devices, including tips
and other essential parts needed for delivering a specified volume, which perform liquid handling tasks
without human intervention into labware.
NOTE For terminology and general requirements of automated liquid handling systems, see ISO 23783-1.
Determination, specification, and reporting of volumetric performance of automated liquid handling systems is
described in ISO 23783-3.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
ISO 3696, Water for analytical laboratory use — Specification and test methods
ISO 8655-6, Piston-operated volumetric apparatus – Part 6: Gravimetric reference measurement procedure
for the determination of volume
ISO 23783-1, Automated liquid handling systems — Part 1: Terminology and general requirements
ISO 23783-3, Automated liquid handling systems — Part 3: Determination, specification, and reporting of
volumetric performance
3 Terms and definitions
For the purposes of this document, the terms and definitions given in ISO 23783-1 apply.
ISO and IEC maintain terminology databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at https:// www .iso .org/ obp
— IEC Electropedia: available at https:// www .electropedia .org/
4 Abbreviated terms
For the purposes of this document, the abbreviated terms given in ISO 23783-1 apply.
ISO 23783-2:2022(E)
5 Measurement methods
5.1 Overview of methods suitable for measuring ALHS performance
When choosing a test method for an ALHS, its suitability for the specific test situation shall be evaluated.
This evaluation shall consider the systematic and random error requirements of the ALHS to which the
test method is being applied. The selected test method shall be adequate to evaluate whether the ALHS
performance is fit for its intended purpose.
NOTE 1 Fitness for purpose is a foundational concept and closely related to the process of metrological
confirmation as described in ISO 9000 and ISO 9001.
The test method shall have a sufficiently small measuring system uncertainty (MSU) for the specific
test situation. The MSU should be determined in accordance with a suitable approach (see 8.2 for more
detail).
NOTE 2 The measurement model approach for estimating MSU is described in ISO/IEC Guide 98-3 and the
[4]
measurement system approach is described in EURACHEM/CITAG Guide CG 4 .
Table 1 is intended to provide an overview of methods suitable for determining the volumetric
performance of ALHS. It provides cross-references between the method abstracts from 5.2 to 5.5, and
the corresponding procedures in Annexes B to I. It further describes the volume ranges, plate and
liquid types which can be used for testing ALHS performance with a given method. It also lists typical
systematic and random errors achievable if a test procedure is exactly followed as described in its
respective annex. The suitability of a method for a given test situation may also be determined by the
required equipment or environmental conditions under which it needs to be carried out.
Only key test equipment is listed in Table 1, while test equipment to monitor liquid and air temperatures,
relative humidity, and barometric pressure is required for each procedure, as specified in the
corresponding annexes.
ISO 23783-2:2022(E)
Table 1 — Test methods for ALHS
Typical Typical
Method Liquid Environmental
Method Plate type Volume range systematic random Test equipment
b
Ref. type conditions
a a
error error
wells µl % %
Photometric methods
5.2.1 Dual-dye ratiom- Aqueous, 96 0,1 to 350,0 2,0 to 3,0 0,15 to 0,25 Temperature: — Microplate absorbance reader capable
c
etric photometric DMSO of measuring absorbance at 520 nm
384 0,01 to 55,0 2,5 to 5,5 0,35 to 0,55
Annex B Aqueous:
method and 730 nm;
15 °C to 30 °C
— dimensionally characterized 96- or
c
DMSO :
384-well microplates with optically
clear bottom;
19 °C to 30 °C
d
RH : 20 % to 90 %
— calibration plate for plate reader;
— microplate shaker;
— balance;
— spectrophotometer capable of
measuring absorbance at 520 nm and
730 nm;
— pH meter;
— volumetric flasks.
a
Typically, larger test volumes lead to smaller errors.
b
The minimum temperature of the test environment shall be above the melting point of the test liquid, ensuring that it will not solidify at any point during the test. The relative
humidity of the test environment shall be non-condensing.
c
Dimethylsulfoxide.
d
Relative humidity.
ISO 23783-2:2022(E)
Table 1 (continued)
Typical Typical
Method Liquid Environmental
Method Plate type Volume range systematic random Test equipment
b
Ref. type conditions
a a
error error
wells µl % %
5.2.2 Single-dye photo- Aqueous 96 1,0 to 100,0 3 1,5 Temperature: — Microplate absorbance reader capable
metric method of measuring absorbance at 492 nm
384 0,25 to 20,0 3 1,5
Annex C 15 °C to 30 °C
and 620 nm;
d
RH : 40 % to 70 %
— 96- or 384-well microplates with
optically clear bottom;
— balance;
— magnetic stirrer;
— microplate shaker;
— pH meter;
— manual pipettes;
— volumetric flasks.
a
Typically, larger test volumes lead to smaller errors.
b
The minimum temperature of the test environment shall be above the melting point of the test liquid, ensuring that it will not solidify at any point during the test. The relative
humidity of the test environment shall be non-condensing.
c
Dimethylsulfoxide.
d
Relative humidity.
ISO 23783-2:2022(E)
Table 1 (continued)
Typical Typical
Method Liquid Environmental
Method Plate type Volume range systematic random Test equipment
b
Ref. type conditions
a a
error error
wells µl % %
5.2.3 Fluorescence meth- Aqueous, 384 0,001 to 0,015 <8 <8 Temperature: — Microplate fluorescence reader with
od excitation wavelength at 494 nm and
c
1 536 0,001 to 0,015 <8 <8
Annex H DMSO 17 °C to 27 °C
emission analysis at 521 nm;
d
RH : non-condensing
— 384- or 1536-well fluorescence
microplates;
— balance;
— bulk liquid dispenser or multi-channel
pipette;
— microplate shaker;
— pH meter;
— manual pipettes;
— volumetric flasks.
a
Typically, larger test volumes lead to smaller errors.
b
The minimum temperature of the test environment shall be above the melting point of the test liquid, ensuring that it will not solidify at any point during the test. The relative
humidity of the test environment shall be non-condensing.
c
Dimethylsulfoxide.
d
Relative humidity.
ISO 23783-2:2022(E)
Table 1 (continued)
Typical Typical
Method Liquid Environmental
Method Plate type Volume range systematic random Test equipment
b
Ref. type conditions
a a
error error
wells µl % %
Gravimetric methods
5.3.1 Single channel anal- Any n/a 0,5 to <20 ≤1,4 ≤0,6 Temperature: — Balance;
ysis
20 to <200 ≤0,9 ≤0,3
Annex D 17 °C to 30 °C
— density meter;
200 to 1 000 ≤0,9 ≤0,3 d
RH : 45 % to 80 %
— anti-electrostatic equipment;
— anti-vibration table;
— temperature- and humidity control for
environment;
— draft shield or draft-free environment
for balance.
5.3.2 Regression analysis Any n/a <0,015 20 to 50 <10 Temperature: — Balance;
0,015 to <0,050 2 to 5 2,5 to 5
Annex E 17 °C to 27 °C
— density meter with 6 decimal places;
0,050 to 1 0,5 to 2 <0,5 d
RH : 45 % to 80 %
— anti-electrostatic equipment;
Barometric pressure:
— anti-vibration table;
600 hPa to 1 100 hPa
— temperature- and humidity control for
environment;
— draft shield or draft-free environment
for balance.
a
Typically, larger test volumes lead to smaller errors.
b
The minimum temperature of the test environment shall be above the melting point of the test liquid, ensuring that it will not solidify at any point during the test. The relative
humidity of the test environment shall be non-condensing.
c
Dimethylsulfoxide.
d
Relative humidity.
ISO 23783-2:2022(E)
Table 1 (continued)
Typical Typical
Method Liquid Environmental
Method Plate type Volume range systematic random Test equipment
b
Ref. type conditions
a a
error error
wells µl % %
Photometric/gravimetric hybrid method
5.4 Tartrazine as Aqueous 96 1,0 to 300,0 0,2 to 0,8 0,5 to 1,0 Temperature: — Balance;
chromophore
Annex F
— microplate absorbance reader
capable to measure absorbance at the
following wavelengths, depending on
the chromophore used:
384 1,0 to 20,0 0,4 to 1,0 0,9 to 1,5 17 °C to 30 °C — 4-nitrophenol: 405 nm and 620 nm,
4-nitrophenol as Aqueous 96 10 to 1 000 <1 to 5 1 to 2 Temperature stability: — Tartrazine: 450 nm and 620 nm,
chromophore
5 to 250 < ±0,5 °C
d
96, 384 1 to 60 <1 to 5 1 to 2 RH : 45 % to 80 % — Orange G: 492 nm and 620 nm
d
0,5 to 25 RH stability:
< ±10 %
384, 1 536 0,1 to 8 <2 to 10 2 to 5 — microplate shaker;
— 96- or 384-well microplates with
optically clear bottom;
— manual pipettes;
— centrifuge tubes 1,5 ml;
— anti-vibration table;
Orange G as chromo- Aqueous 96 1 to 100 <1 to 5 1,5 — temperature- and humidity control for
phore environment;
384 1 to 50 <1 to 5 1,5 — draft shield or draft-free environment
for balance.
a
Typically, larger test volumes lead to smaller errors.
b
The minimum temperature of the test environment shall be above the melting point of the test liquid, ensuring that it will not solidify at any point during the test. The relative
humidity of the test environment shall be non-condensing.
c
Dimethylsulfoxide.
d
Relative humidity.
ISO 23783-2:2022(E)
Table 1 (continued)
Typical Typical
Method Liquid Environmental
Method Plate type Volume range systematic random Test equipment
b
Ref. type conditions
a a
error error
wells µl % %
Dimensional methods
5.5.1 Optical image anal- Any n/a Free flying drop- <5 <2 Temperature: — Stroboscopic camera or high-speed
ysis of droplets lets of V <0,5 µl camera;
Annex G (20 ± 3) °C or (27 ± 3) °C
d
— automatic image detection software.
RH : 50 % to 80 %
5.5.2 Optical image analy- Any n/a 0,1 to 1,0 <10 <7 Temperature: — Flatbed scanner;
sis of capillaries
>1,0 to 1 000 <5 <4
Annex I 15 °C to 35 °C
— image analysis software;
d
RH : 15 % to 90 %
— specialized plates with capillaries.
a
Typically, larger test volumes lead to smaller errors.
b
The minimum temperature of the test environment shall be above the melting point of the test liquid, ensuring that it will not solidify at any point during the test. The relative
humidity of the test environment shall be non-condensing.
c
Dimethylsulfoxide.
d
Relative humidity.
ISO 23783-2:2022(E)
5.2 Photometric methods
5.2.1 Dual-dye ratiometric photometric method
This method allows the determination of volumes of aqueous test liquids from 0,1 µl to 350 µl in 96-
well plates, and from 0,01 µl to 55 µl in 384-well plates. Volumes of dimethylsulfoxide (DMSO)-based
test liquids can be determined from 0,11 µl to 10 µl in 96-well plates, and from 0,01 µl to 2,5 µl in 384-
well plates.
This method is suitable to determine the performance of ALHS with up to 384 channels. The operating
environment for this method is 15 °C to 30 °C (19 °C to 30 °C for DMSO liquids), and it is not dependent
on the ambient relative humidity and barometric pressure at the test location. Further information on
the effect of relative humidity and barometric pressure on this method can be found in Reference [5].
Traceability of the measurement results to the International System of Units (SI) is achieved through the
use of a calibrated microplate absorbance reader, dimensionally characterized microplates, calibrated
balance, and calibrated volumetric glassware.
The procedure for the dual-dye ratiometric photometric method specified in Annex B shall be followed.
5.2.2 Single-dye photometric method
This method is suitable for evaluating the volumetric performance of ALHS with up to 384 channels
using aqueous test liquids. Volumes from 1 µl to 100 µl can be measured in 96-well plates, and from
0,25 µl to 20 µl in 384-well plates.
Traceability of the measurement results to the SI is achieved through the use of a calibrated balance,
calibrated pipettes, a calibrated microplate absorbance reader, and calibrated volumetric glassware.
The procedure for the single-dye photometric method specified in Annex C shall be followed.
5.2.3 Fluorescence method
This method is suitable to evaluate the volumetric performance of ALHS delivering volumes smaller
than 15 nl. The fluorescence of the test liquid of fluorescein in DMSO is measured in 384-well or 1536-
well microplates, which are specifically suited for fluorescence measurements.
This method is intended to be used for non-contact liquid delivery devices (e.g. acoustic, dispensing
valves, or inkjet-type technology) that deliver the liquid volume as free flying droplets or jets into the
wells of the microplate.
Traceability of the measurement results to the SI is achieved through the use of a calibrated fluorescence
microplate reader, calibrated balance, calibrated pipettes, and calibrated volumetric glassware.
The procedure for the fluorescence method specified in Annex H shall be followed.
5.3 Gravimetric methods
5.3.1 Single channel method
This method describes the apparatus, procedure and reference material for recording measurements
with the gravimetric method. A single pan balance is used to take a measurement from a single channel
at a time. The following accommodations shall be made:
— placement of the balance and the weighing vessel which reduce draft and vibrations to a suitable
level;
— control of the environmental conditions affecting the mass to volume conversion of the measurement
(temperature and relative humidity);
ISO 23783-2:2022(E)
— monitoring of the barometric pressure, which affects the mass to volume conversion.
Traceability of the measurement results to the SI is achieved through the use of a calibrated balance
and accounting for test liquid density and air buoyancy.
The procedure for the single-channel gravimetric method specified in Annex D shall be followed.
5.3.2 Regression analysis
The gravimetric regression method (GRM) is suitable for the measurement of very small liquid volumes,
between 0,005 µl and 1 µl. The method is based on a gravimetric balance as the primary measurement
device.
This method is intended to be used for non-contact liquid delivery devices (e.g. dispensing valves,
acoustic, or inkjet-type dispensing) that deliver the liquid volume as free flying droplets or jets to the
balance receptacle.
The key difference to traditional gravimetric methods used for the measurement of larger volumes is
the determination of the target volume: a series of balance readings is recorded over a period of time
before and after the device under test has delivered the liquid to be measured into the receptacle on
the balance. The measurement result of the delivered test liquid is then determined as the difference
between two linear regression lines fitted to the recorded balance data before and after the liquid
delivery. This method allows measurement of balance drift due to evaporation and other disturbances
of the measurement (e.g. by vibrations during the data acquisition), so that these can be compensated
for in the measurement calculation (see Reference [6] for more details).
Accommodations regarding the placement of the balance and environmental control and monitoring
given in 5.3.1 shall be made.
Traceability of the measurement results to the SI is achieved through the use of a calibrated balance.
The procedure for the gravimetric regression method specified in Annex E shall be followed.
5.4 Hybrid photometric/gravimetric method
The photometric / gravimetric hybrid method allows the evaluation of volumetric performance of ALHS
by a combination of a gravimetric measurement with subsequent photometric measurements. Test
liquid containing a chromophore is delivered into 96-well or 384-well microplates. The systematic error
is determined by gravimetry of the aggregate deliveries into the microplate. Subsequently, the random
error of volume deliveries is determined photometrically by measuring the relative absorbances of each
well of the microplate.
Chromophores suitable for this method are Tartrazine, Orange G, and 4-nitrophenol. The procedure
described in Annex F is suitable for test volumes between 1 µl and 200 µl in 96-well plates, and 1 µl and
50 µl in 384-well plates.
Accommodations regarding the placement of the balance and environmental control and monitoring
given in 5.3.1 shall be made.
Traceability of the measurement results to the SI is achieved through the use of a calibrated balance,
calibrated pipettes, and calibrated volumetric glassware.
The procedure for the hybrid method specified in Annex F shall be followed.
5.5 Dimensional methods
5.5.1 Optical image analysis of droplets
This method measures the volume of delivered liquids by analysing images acquired by a high-
speed camera and stroboscopic illumination during the liquid delivery cycle. It is suitable for ALHS,
ISO 23783-2:2022(E)
which deliver liquid volumes as a sequence of discreet micro droplets (for further information on the
determination of droplet volumes, see Reference [7]).
Traceability of the measurement results to the SI is achieved through calibration of the length scale of
the acquired images.
The procedure for the optical image analysis of droplets specified in Annex G shall be followed.
5.5.2 Optical image analysis of capillaries
The method is based on the optical analysis of images acquired of capillaries of known and calibrated
geometry. For the image acquisition, a flatbed scanner is used. The method provides a direct
determination of the volume by the optical measurement of one or multiple lengths of the calibrated
capillaries.
The method can be used to measure volumes between 0,1 µl and 1 000 µl for ALHS with 1 to 384
channels. The measurement uncertainties only minimally depend on the environmental conditions and
are reliable in a broad range of environmental conditions without error corrections.
Traceability of the measurement results to the SI is achieved by using calibrated capillaries and a
calibrated image acquisition device.
The procedure for the optical image analysis of capillaries specified in Annex I shall be followed.
6 Equipment and preparation
6.1 Test equipment
Test equipment used for volumetric performance measurements according to the procedures described
in this document shall conform to the minimum performance requirements given in Tables 2, 3, 4, and
5, unless different minimum performance requirements are described within a specific test procedure.
Balances shall be allowed to settle for at least 6 s before reading the indicated value.
NOTE 1 The balance repeatability and expanded uncertainty in use given in Table 2 are harmonized with
Table 3 but the weighing ranges are restricted to improve accuracy when weighing an amount of dry chemicals.
NOTE 2 The balance requirements given in Table 3 are based on ISO 8655-6 and allow multiple replicate
deliveries of test liquid into the same weighing vessel without emptying it out.
Table 2 — Minimum requirements for balances for weighing dry materials
Smallest amount to be Expanded uncertainty in
Readability Repeatability
a
weighed use (k = 2)
g mg mg mg
<1,0 0,001 0,006 0,012
1,0 0,01 0,025 0,05
10 0,1 0,2 0,4
100 1 2 4
1 000 10 20 40
a [8] [9]
Uncertainty in use can be determined according to ASTM E898-20 and EURAMET CG-18 at the minimum mass
listed in the table.
ISO 23783-2:2022(E)
Table 3 — Minimum requirements for balances for weighing liquids
Delivered volume of test Expanded uncertainty in
Readability Repeatability
a b
liquid use (k = 2)
µl mg mg mg
<0,5 0,000 1 0,000 5 0,001
0,5 ≤ V < 20 0,001 0,006 0,012
20 ≤ V < 200 0,01 0,025 0,05
200 ≤ V ≤ 10 000 0,1 0,2 0,4
a
Assumes one delivery of test liquid from a single channel.
b [8] [9]
Uncertainty in use can be determined according to ASTM E898-20 and EURAMET CG-18 at the value of the largest
volume of the listed range, assuming a single channel delivery.
Table 4 — Minimum performance requirements for absorbance microplate readers
Parameter Requirement
b
Photometric resolution 0,001 AU
a
Photometric trueness from 0 AU to 1,0 AU 0,005 AU
a
Photometric trueness from > 1,0 AU to 2,0 AU 0,010 AU
Photometric repeatability from 0 AU to 2,0 AU 0,005 AU
System linearity between 0 AU and 2,0 AU 0,010 AU
Wavelength accuracy < ± 1,5 nm
a
Photometric trueness is sometimes called photometric accuracy.
b
Absorbance unit.
Table 5 — Minimum performance requirements for other test equipment
Instrument Resolution Expanded uncertainty
(k = 2)
Thermometer for liquids 0,1 °C 0,2 °C
Thermometer for ambient air 0,1 °C 0,2 °C
a a
Hygrometer 1 % RH 5 % RH
Barometer 0,1 kPa 1 kPa
Timing device 1 s not relevant
a
Relative humidity.
6.2 Manually operated single- and multi-channel pipettes
Single-channel and multi-channel pipettes shall be calibrated and shall fulfil the minimum performance
requirements given in ISO 8655-2. Operator impact on manually pipetted volumes shall be considered
in the calculation of errors and measurement uncertainty.
6.3 Preparation for testing
The ALHS under test, including all exchangeable parts to be used during the test, and all test equipment
and test liquids shall be in thermal equilibrium (±2 °C) for at least 2 h prior to the start of testing. During
the time of testing, the environmental conditions shall not change more than ±1 °C and 5 % relative
humidity (RH). Requirements for environmental conditions are described in each test procedure.
Laboratory instrumentation is designed to be operated under non-condensing humidity conditions.
All test equipment shall be calibrated according to the test procedure in this document, or according to
the manufacturer’s instructions if the calibration is not specifically explained in the procedure.
ISO 23783-2:2022(E)
The ALHS under test and test equipment shall be powered on and allowed sufficient time to equilibrate
for proper functionality according to the manufacturer’s instructions.
Liquid reservoirs of ALHS shall be filled immediately before testing begins. ALHS which require priming
shall be primed according to the manufacturer’s instructions immediately before testing begins.
The tips of piston-operated ALHS should be pre-rinsed with the test liquid at least five times, whereby
the dispensed test liquid is discarded to waste. This step is required every time a tip is changed.
7 Thermal expansion
If the test temperature is different from the temperature of adjustment of the ALHS, and if the cubic
thermal expansion coefficient γ of the volumetric apparatus is known, Formula (1) may be used to
correct the measured volume of test liquid for thermal expansion:
VV=× 1−×γ ()tt− (1)
[]
L,tc LT ref
where
V is the delivered volume of test liquid, corrected for thermal expansion of the ALHS;
L,tc
V is the delivered volume of test liquid measured at the test temperature;
L
γ is the cubic thermal expansion coefficient of the volumetric apparatus;
t is the temperature at which the test is performed;
T
t is the reference temperature of adjustment of the ALHS.
ref
8 Traceability and measuring system uncertainty
8.1 Traceability
All measurements by relevant test equipment used for the calibration of an ALHS shall be traceable to
the International System of Units (SI). The uncertainty of relevant test equipment affects the error of
the reported volumetric results.
NOTE Relevant test equipment can include but is not limited to: balances, thermometers, pH meters,
hygrometers, plate readers, spectrophotometers, calibrated pipettes, volumetric glassware, size calibration
charts, and timing devices.
8.2 Estimation of measuring system uncertainty
8.2.1 Whole system approach
Measuring system uncertainty (MSU) may be estimated by statistical evaluation of results produced
by the entire measuring system. Precision and bias studies, measurement system analysis, and inter-
laboratory comparisons are some of the means by which this is achieved. Detailed approaches are
described in Reference [4].
8.2.2 Measurement model approach
This approach estimates MSU based on an analysis of each input to a measurement model. This approach
is detailed in Reference [3].
ISO 23783-2:2022(E)
9 Reporting
Measurement results, traceability, and measurement uncertainty shall be reported in accordance with
ISO 23783-3.
ISO 23783-2:2022(E)
Annex A
(normative)
Calculation of liquid volumes from balance readings
A.1 Calculation of liquid volume from the balance reading
A.1.1 General formula for volume
For the conversion of the balance reading of the mass, m, to volume, V, at the test temperature, a
correction for the liquid’s density and air buoyancy is necessary. The calculation of the liquid volume at
the test temperature is given by Formula (A.1).
In case the test liquid is water and the calculations given in Clause A.1 are not
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