Instructions for Use

Absorbance 96 App
Version 1.2

Instructions for Use

Absorbance 96 App
Version 1.0 (18/01/2021)
©Copyright 2021 by Byonoy GmbH. All rights reserved.

Manufacturer: Byonoy GmbH 00 49 40 5379 866 00

Bernstorffstr. 118 [email protected]
22767 Hamburg, Germany www.byonoy.com
TABLE OF CONTENTS

1 General information ...................................................................................................................... 1

2 App installation procedure and updates ........................................................................ 2

3 Absorbance 96 App overview ............................................................................................... 3

4 Operation............................................................................................................................................ 5

4.1 Set up an assay and create a protocol ........................................................................................ 5

4.2 Readout of the assay ................................................................................................................................ 8
4.3 Results.................................................................................................................................................................. 8

5 EC-Conformity Declaration ..................................................................................................... 9

II
 General information

1 General information

The Absorbance 96 App is to be used only in combination with the Absorbance 96 reader.
The Absorbance 96 App is not a stand-alone product. Please also refer to the separate op-
erating instructions for the Absorbance 96 reader.

The Absorbance 96 App is only compatible with the Absorbance 96 reader. The safeties,
precautions and validations included in the Absorbance 96 App have only been tested for
the Absorbance 96 and shall not be used for any other purpose on any other device.

In this manual, particularly important remarks are labeled as followed:

Warning:
This is a warning that indicates an immediately or potentially dangerous situa-
tion, which, if not avoided, will result in death or serious injury.

Caution:
Caution indicates a potentially hazardous situation, which, if not avoided, could
result in minor or moderate injury.

Information:
This is a piece of information indicating certain properties that must be ob-
served.

About this manual

Important: Follow the instructions for use

This user manual may only be used according to its intended purpose. It may not be repro-
duced, changed, or translated in another language without the prior written consent of
Byonoy GmbH.

This document is subject to technical changes and updates.

Ensure that the manual is always accessible and is read and understood by all
persons operating the instrument.

The latest version of this manual is available via the Absorbance 96 App.

1
 App installation procedure and updates

2 App installation procedure and updates

The Absorbance 96 App is compatible with Microsoft Windows (Windows 7 or above) and
Mac OS (High Sierra 10.13. or above).

To install the Absorbance 96 App, insert the USB stick into the computer and follow the steps
below:

• Locate and select the USB drive in the document browser.

• Open the installation file, either for Windows or macOS.
• Follow the steps in the Setup Wizard to complete the installation.

Modern operating systems offer plug and play functionality. Older operating systems may
require manual installation of the software drivers. The drivers for Windows are available on
the USB stick.

To install the proper drivers and software on the computer, the user must have
administrator rights or obtain such permissions.

Uninstalling the software

The Absorbance 96 App can be uninstalled using the computer’s standard uninstall proce-
dure.

Updates

To check for new updates, click the update button in the lower left corner. If an update is
available, this will be shown in a bar at the top of the screen.

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 Absorbance 96 App overview

3 Absorbance 96 App overview

The Absorbance 96 App is used to operate the Absorbance 96 reader and process the re-
sults. After the app is started and the reader is connected, the main window appears auto-
matically.

Figure 1. Main window.

Icons in the leftmost menu bar (from top to bottom):

1. New: Start creating a protocol and/or assay.

2. Open: Open a previously saved protocol or assay.
3. Save As: Save a current protocol or assay.
4. Manual: Open the app manual.
5. Update: Update the app.

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 Absorbance 96 App overview

File Types

The following file types are used by the Absorbance 96 App:

Table 1. File Types in the Absorbance 96 App

Description Extension
Protocol .byop
Assay .byoa
Export .csv/.pdf

The protocol file type .byop allows to save settings and to load predefined protocols for re-
peat assays.

The assay file type .byoa is used to save completed assays. In addition to the settings, the
results of the assay are also saved.

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 Operation

4 Operation

4.1 Set up an assay and create a protocol

Select “New” from the menu bar. Your experiment conditions can now be defined.

Workflow Parameters

1. Wavelengths
Under “Wavelengths”, a signal and reference wavelength (both in nanometers) may be cho-
sen.

The signal wavelength is the wavelength at which the principal measurement will be run.

The reference wavelength is an additional wavelength at which the device will measure and
whose results will be subtracted from the values measured at the signal wavelength. If no
reference wavelength is desired, click the “None” button beside the reference wavelength.

2. Readout Method
Select your readout method, either Endpoint or Kinetic.

The kinetic readout method is not available for every reader type.

To perform kinetic measurements and avoid misinterpretation of the results, observe the
following instructions:

Ensure constant environmental conditions, i. e. constant ambient temperature

and humidity.

Acclimatize the connected reader and microtiter plate to the ambient tempera-
ture for at least 1 hour.

A signal drift of about 1 mOD per hour might be observed.

3. Evaluation
The evaluation section allows to choose which evaluative method will be used for the
readout.

No Evaluation

No evaluation will be used. Only the OD values will be shown.

Quantitative

Upon selection of the quantitative evaluation method, a drop-down menu will appear. Here,
a fitting method can be chosen. The selected fitting method will be used for the standard
curve and the data analysis.

5
 Operation

Predefined standard curve

In a quantitative assay, a predefined standard curve can be used to determine the concen-
trations of a new set of samples with identical experiment parameters. After selecting the
checkbox “Use predefined standard curve”, you may enter the curve parameters manually
or load a standard curve from a previous assay.

After selecting the checkbox “Use predefined standard curve”, you cannot as-
sign any new standards.

In a quantitative assay, the standard curve is a function of various assay pa-

rameters, such as the temperature, timing, or age of the assay kit. Before using
this feature, it is advisable to validate your assay for compatibility.

As the above-mentioned experiment conditions are subject to change, it is rec-

ommended to use a new standard curve when performing a new assay.

Qualitative

Upon selection of qualitative evaluation, a formula for the cut-off definition will appear.

Figure 2. Formula for cut-off definition.

Terms for cut-off definition

The terms for cut-off definition from left to right according to Figure 2:

• POS = the mean of the OD values of the positive control wells.

• NEG = the mean of the OD value of the negative control wells.
• CUT = the mean of the OD value of the cut-off control wells.
• 0 OD = zero. User-defined value may be entered.
• Offset = an assay dependent factor provided by the user.
• Range = definition of a range as a percentage value.

The formula setup for the cut-off definition covers multiple formula types. These are listed
below.

• Cut-Off Value = mean negative controls +(-) k

• Cut-Off Value = mean positive controls + k
• Cut-Off Value = mean cut-off controls + k
• Cut-Off Range = [mean control + k] +(-) x %

k is an experimentally determined factor normally defined by the laboratory professional or

by the kit manufacturer.

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 Operation

Example:

To replace the minus function, the user must simply enter a negative value. For example, if
the cut-off is

Cut-off = Mean Negative Controls - 0.2,

this must be entered as follows:

Figure 3. Cut-off calculation with subtraction

In case the Cut-Off is known or provided by the kit manufacturer, and therefore does not
need to be calculated, select 0 OD and enter the value into the middle field:

Figure 4. Entering a known cut-off value

The examples shown in this manual do not represent actual data, rather func-
tion to clarify software features, and may not be referenced for experimental
methods or procedure.

Plate Layout

In the plate layout section, samples, standards, blanks, and controls can be defined. Select
the type from the options below the plate layout; click and drag to define. Duplication pat-
terns allow the user to define multiple replicates for samples and standards.

The format of the plate layout and the corresponding buttons are defined by
the evaluation method. Quantitative and qualitative evaluations have different
fields and buttons.

When replicates are defined, the mean value is used for the calculations.

In the plate layout for a quantitative assay, a field is provided where the concentrations of
the standards and the samples can be entered. The unit can be entered manually or chosen
from the dropdown menu. Using the dilution factor, diluted samples can be used, and the
software will calculate the undiluted concentration.

B0 control

B0 is a specific denomination of zero standard controls in competitive ELISAs. If B0 is chosen,

its value is used to reference the relative binding in a sample or standard to the maximum
possible binding. This ratio is referred to as the percentage of binding %B0.

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 Operation

Create a protocol

Once the parameters of a given experiment have been set, these settings may be saved as
a protocol. All parameters, such as wavelength and plate layout, will be saved.

4.2 Readout of the assay

Once all settings have been completed, the read out can be started. The app will automati-
cally guide the user through the readout process.

4.3 Results

After a successful measurement, the results section will open automatically. The results are
displayed in different views:

Plate view: Provides an overview of the different OD values in their respective wells. Unde-
fined wells are not shown. Click on a given well to see the OD value, label, and, if applicable,
concentration.

Graph view: A representation of the data will be shown on the graph. The standard curve is
dependent on the fitting method chosen. By choosing the right fitting method, the distance
between the curve and each point is minimized. Using the standard curve, the concentration
of each sample is calculated. The equation below the graph reflects the fitting method ap-
plied and offers further data.

Tabular view: Provides an overview of the various samples, wells, OD values, and calculations.

Table 2. Overview of terms, tabular view section

Parameter Description

ID The assigned ID of a given standard

Well The assigned well

Absorbance (OD) The absorbance reading in OD units

The coefficient of variation between repli-
Concentration %CV
cates
The entered concentration values for the
Standard Concentration
standards
Concentration The calculated concentration values

Mean Absorbance (OD) The mean OD value of replicate wells

Pos/Neg/Intermediate result of a qualita-
Result*
tive experiment
(*Only appears for qualitative evaluations)

Export

Export the results as a .pdf or .csv file.

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 EC-Conformity Declaration

5 EC-Conformity Declaration