Walz MINI-PAM-II User manual

2.163/04.2018
3. Edition, May 2018
MINI_PAM_II_03.docx
Heinz Walz GmbH, 2018
Heinz Walz GmbH •Eichenring 6 •91090 Effeltrich •Germany
Phone [phone] •Telefax +49-(0)9133/5395
E-mail [email protected]om •Internet www.walz.com
MINI-PAM-II
Manual for
Standalone Use

Contents Chapter 1
C1
1 Contents
1 Contents .............................................................. 1
2 Safety Instructions.............................................. 1
2.1 General Safety Instructions ............................................ 1
2.2 Special Safety Instructions.............................................. 1
3 Introduction ......................................................... 3
3.1 Overview............................................................................... 4
4 Components and Setup ...................................... 7
4.1 Extent of Delivery (Basic System) ................................ 7
4.2 Optoelectronic Unit............................................................ 8
4.2.1 Batteries.............................................................................. 11
4.3 2010-A Distance Clip 60° .............................................. 11
4.4 Accessories........................................................................ 13
4.4.1 2035-B Leaf-Clip Holder ................................................ 13
4.4.2 DLC-8 Dark Leaf Clip...................................................... 16
4.4.3 2060-B Arabidopsis Leaf Clip....................................... 18
4.4.4 2060-A Fiberoptics Holder for Surfaces.................... 18
4.4.5 2065-M Mini Quantum/Temp.-Sensor....................... 19
4.4.6 2054-L External LED Source ....................................... 20
4.4.7 MINI-SPEC/MP Miniature Spectrometer .................. 22
4.4.8 KS-2500 Suspension Cuvette...................................... 27
4.4.9 Oxygen Package.............................................................. 27
4.4.10 BCS-9590 Barcode Scanner........................................ 28
4.4.11 MINI-PAM/F1 Miniature Fiberoptics........................... 29
4.4.12 MQS-B/A Adapter Set for Thin Fiberoptics.............. 30
5 Saturation Pulse Analysis ................................ 31
5.1 Five Fluorescence Levels.............................................. 31
5.1.1 Measurements with Dark-Acclimated Samples...... 32
5.1.2 Measurements with Illuminated Samples................. 32

Chapter 1 Contents
C2
5.2 Fluorescence Ratio Parameters ..................................36
5.3 Relative Electron Transfer Rate (ETR)......................38
5.4 Light Curves .......................................................................39
5.5 Some Light Curve References .....................................42
5.6 Some Reviews on Saturation Pulse Analysis..........43
6 Hints & Troubleshooting ...................................45
6.1Instrument Settings..........................................................45
6.2 Default settings .................................................................45
6.3 F0Fluorescence ...............................................................46
6.5 Signal Noise .......................................................................48
6.6 System Hangs ...................................................................48
6.7 External PAR Sensor is not Responding ..................48
6.8 Intensity of external lamp cannot be adjusted.........48
7 Touchscreen Operation.....................................49
7.1 Calibration...........................................................................49
7.2 Top Level Windows .........................................................50
7.2.1 Basic Data ..........................................................................52
7.2.2 Primary Data ......................................................................55
7.2.3 Quenching Analysis.........................................................56
7.2.4 Ft-Chart................................................................................57
7.2.5 Spectrometer .....................................................................58
7.2.6 Actinic + Yield ....................................................................60
7.2.7 Induction Curve.................................................................61
7.2.8 Light Curve .........................................................................64
7.2.9 Recovery .............................................................................64
7.2.10 Actinic Light List ................................................................66
7.3Main Menu..........................................................................70
7.3.1 PAM Settings .....................................................................70
7.3.1.1 Meas. Light.........................................................................71
7.3.1.2 Meas. Light Sett................................................................72
7.3.1.3 Gain ......................................................................................74

Contents Chapter 1
C3
7.3.1.4 Damping.............................................................................. 74
7.3.1.5 ETR-Factor ........................................................................ 75
7.3.1.6 Fo’ Mode............................................................................. 75
7.3.1.7 Adjust F-Offset.................................................................. 75
7.3.2 Light Sources .................................................................... 77
7.3.2.1 Far Red Sett. ..................................................................... 78
7.3.2.2 Light Panel Sett................................................................ 78
7.3.2.3 SAT Settings...................................................................... 79
7.3.3 Program/Clock Settings ................................................. 80
7.3.3.1 Actinic + Yield.................................................................... 81
7.3.3.2 Induction Curve ................................................................ 82
7.3.3.3 Light Curve......................................................................... 83
7.3.4 Sensors ............................................................................... 85
7.3.4.1 Internal PAR ...................................................................... 86
7.3.4.2 Leaf Clip/Ext. PAR........................................................... 88
7.3.4.3 Oxygen Sensor................................................................. 92
7.3.4.4 Spectrometer..................................................................... 93
7.3.4.5 Load System Settings..................................................... 96
7.3.5 MINI-PAM-II Settings...................................................... 96
7.3.6 Memory ............................................................................. 101
7.3.7 Info...................................................................................... 101
7.3.7.1 MINI-PAM-II Info............................................................. 102
7.3.7.2 Sensor Info...................................................................... 103
7.3.7.3 Firmware Info ................................................................. 104
8 Specifications MINI-PAM-II ............................. 105
8.1 Basic System................................................................... 105
8.1.1 Optoelectronic Unit........................................................ 105
8.1.2 Fiberoptics MINI-PAM/F .............................................. 107
8.1.3 Power Supply MINI-PAM-II/N..................................... 107
8.1.4 Battery Charger 000190101101................................ 108
8.1.5 Distance Clip 60° 2010-A ............................................ 108

Chapter 1 Contents
C4
8.1.6 Complementary Items...................................................108
8.1.7 Software WinControl-3..................................................108
8.1.8 Transport Case MINI-PAM/T ......................................109
8.2 Accessories......................................................................109
8.2.1 2035-B Leaf-Clip Holder...............................................109
8.2.2 Fiberoptics Adapter 90° 2030-B90............................110
8.2.3 2054-L External LED Source......................................110
8.2.4 Dark Leaf Clip DLC-8 ....................................................111
8.2.5 Arabidopsis Leaf Clip 2060-B.....................................111
8.2.6 Fiberoptics Holder for Surfaces 2060-A..................112
8.2.7 Mini Quantum/Temp.-Sensor 2065-M .....................112
8.2.8 Miniature Spectrometer MINI-SPEC/MP.................113
8.2.8.1 Flat Entrance Optics SPEC/P.....................................113
8.2.8.2 Fluorescence and Reflection Optics SPEC/R .......113
8.2.8.3 PAR Calibration Block 000160101439 ....................114
8.2.9 Suspension Cuvette KS-2500 ....................................114
8.2.10 Magnetic Stirrer with Fiberoptics Holder MKS-2500
..............................................................................................115
8.2.11 Compact Tripod ST-2101A..........................................115
8.2.12 MINI-PAM/F1 Miniature Fiberoptics .........................115
9 Warranty ...........................................................117
9.1 Conditions.........................................................................117
9.2 Instructions .......................................................................118
10 Index .................................................................119

Safety Instructions Chapter 2
1
2 Safety Instructions
2.1 General Safety Instructions
- Read safety instructions and the operating instructions
prior to operation of the device and its accessories.
- Pay attention to all safety warnings.
- Keep device and its accessories away from water or
high moisture areas.
- Keep the device and its accessories away from dust,
sand and dirt.
- Do not put the device and its accessories near sources
of heat.
- Ensure that neither liquids nor foreign bodies get inside
the device or its accessories.
- Ensure sufficient ventilation.
- Connect the device only to the power source indicated
in the operating instructions or on the device. If the de-
vice is not in use, remove the mains plug from the sock-
et.
- The device and its accessories should only be repaired
by qualified personnel.
2.2 Special Safety Instructions
- The MINI-PAM-II is a highly sensitive instrument which
should be only used for research purposes, as specified

Chapter 2 Safety Instructions
2
in this manual. Follow the instructions of this manual in
order to avoid potential harm to the user and damage to
the instrument.
- The MINI-PAM-II can emit very strong light! In order to
avoid harm to your eyes, never look directly into the light
port of the MINI-PAM-II or its fiberoptics.
- Switch off MINI-PAM-II before connecting or disconnec-
tion 2054-L External LED Source.

Introduction Chapter 3
3
3 Introduction
- The “Photosynthesis Yield Analyzer MINI-PAM-II” has been
designed for highly sensitive saturation pulse analysis of
photosystem II (PS II) in the field as well as in the laboratory.
The automatically calculated parameters are FV/FM
(maximum photochemical yield), Y(II) (effective photochemi-
cal yield) and its complementary yields Y(NPQ) and Y(NO),
as well as parameters of photochemical (qL, qP) and non-
photochemical quenching (qN, NPQ) (see Table 3, page 34).
- The instrument continues the tradition of the preceding MINI-
PAM chlorophyll fluorometer. The major technical advance-
ments of the MINI-PAM-II are the consistent use of energy-
efficient LEDs, an internal PAR sensor, and stand-alone op-
eration by a touchscreen which is well readable under natu-
ral light conditions. Also, a far red LED has been added for
selective photosystem I excitation.
- A further technical progress is the newly designed leaf clip
sensor (2035-B) which measures photosynthetically active
radiation (PAR) at leaf level with high accuracy and, thus,
provides reliable light intensity data for calculations of elec-
tron transport rates (ETR).
- A variety of add-ons make the MINI-PAM-II a highly versatile
measuring system which can be configured to meet the
needs of the research goal. The range of accessories in-
cludes a multi-colored lighting unit (Section 4.4.6, page 20),
an optical oxygen sensor (Section 4.4.9, page 27) and a
miniature spectrometer (Section 4.4.7, page 22).
- For long-term field campaigns, the memory capacity has
been upgraded to keep data of more than 27,000 saturation

Chapter 3 Introduction
4
pulse analyses. The fluorometer is powered by of-the-shelf
AA (Mignon) batteries which are easily replaceable even un-
der field conditions. One set of batteries lasts for up to 1000
saturation pulse analyses.
- The MINI-PAM-II can be operated in the stand-alone mode
or by the well-proven WinControl-3 software. WinControl-3
has been introduced with the JUNIOR-PAM fluorometer and
now operates many other fluorometers like the DIVING-
PAM, MONITORING-PAM, and WATER-PAM. In addition to
the features available in the stand-alone mode, the software
allows evaluations of light-response curves by a non-linear
fitting routine and automatic execution of custom-designed
experiments using the built-in batch file feature.
3.1 Overview
The MINI-PAM-II fluorometer provides a vast range of settings
and protocols for measuring fluorescence. To make full use of
these opportunities, it is necessary to become acquainted with
terminology and principles of saturation pulse analysis. There-
fore, the present manual provides a chapter dealing with the ba-
sics of saturation pulse analysis (Chapter 5, page 31).
Chapter 5 also provides a short list of review papers on PAM
chlorophyll fluorescence and saturation pulse analysis (page 43).
Further, a small section is included providing some hints for be-
ginners (Chapter 6, page 45).
In the field, the MINI-PAM-II is mostly operated in the stand-
alone mode by its touchscreen. Chapter 7 (page 49) provides de-
tailed instructions on how to use the touchscreen interface in-
cluding advices on fluorescence induction and light curve pro-
grams.

Introduction Chapter 3
5
Also, this manual includes a section on safe handling of the
MINI-PAM-II (Chapter 2, page 1), and on the extent of delivery of
the basic fluorescence system and its accessories (Chapter 1,
page 7). Further, technical information (Chapter 8, page 105)
and warranty conditions (Chapter 9, page 117) are provided.

Components and Setup Chapter 4
7
4 Components and Setup
4.1 Extent of Delivery (Basic System)
Optoelectronic Unit MINI-PAM-II/B or -/R
Fiber optics MINI-PAM/F
Power Supply MINI-PAM-II/N
Battery charger 000190101101
Battery (12 x) 000160101990
USB cable type A to Mini B 000130606252
Distance Clip 60° 2010-A
Sloped Plexiglas rack 000240313614
Stylus 000160201311
Carrying strap 000150401922
Software WinControl-3
MINI-PAM-II Manual
WinControl-3 Software

Chapter 4 Components and Setup
8
4.2 Optoelectronic Unit
Sockets, fiber optics port and on/off switch of the back side of the
MINI-PAM-II are outlined and numbered in Fig. 1, their properties
and functions are explained in Table 1, page 9.
Fig.
1: Back panel of MINI-PAM-II Power-and-Control-Unit

Components and Setup Chapter 4
9
Table 1: Backside of MINI-PAM-II.
Numbering refers
to Fig. 1, page 8 Function
LEAF CLIP
Socket for 2035-B Leaf-Clip Holder
AUX 1
Electronically configured as LEAF CLIP socket
AUX 2
Electronically configured as LEAF CLIP socket
SYNC
Socket for external light source which emits synchronized with
MINI-PAM-II measuring light
ON/OFF
MINI-PAM-II switch
USB SOCKET
Receptacle for MINI-B USB plug.
EXT. DC
Socket for Power Supply MINI-PAM-II/N
COMP 2
Prepared for peripherals mastering RS232 communication (e.g.
bar code scanner)
COMP 1
Electronically configured as COMP 2
LIGHT PORT
Port for Fiberoptics MINI-PAM/F
Note Great caution should be exercised to prevent dirt or foreign
matter from entering the ports or sockets of the MINI-PAM-II. Do
not force a plug into the wrong socket. Orientate
each plug so that the red dot on the plug coin-
cides with the red dot of the socket. Do not try to
disconnect a plug by pulling at the cable. Dis-
connect plug by pulling at the rippled bushing of the plug.
2
1
3
4
5
6
7
8
9
10

Chapter 4 Components and Setup
10
Two versions of the MINI-PAM-II fluorometer are available. The
MINI-PAM-II/B fluorometer is equipped with a blue LED which is
replaced by a red LED in the MINI-PAM-II/R fluorometer. This
LED is electronically driven to act as measuring and as actinic
light source. In addition, both versions of the MINI-PAM-II offer a
far red LED. Normalized spectra of blue, red and far red LEDs
are shown in Fig. 2.
Fig.
2: Normalized Emission Spectra of MINI-PAM II LEDs. Normalized
emission spectra of blue LED (MINI-PAM-II/B), red LED (MINI-PAM-
II/R) and far red LED (MINI-PAM-II/B and R).

Components and Setup Chapter 4
11
4.2.1 Batteries
When operated independently, the system is powered by six AA
(Mignon) rechargeable batteries (1.2 V/2 Ah). The system can
also be powered by non-rechargeable batteries. The battery
compartment of the opto-electronic unit does not have a charging
function. Therefore, the device can be connected to line power
even in the presence of non-rechargeable batteries.
The battery compartment is closed by an aluminum plate. Its
locking mechanism functions properly if the label “INNER FACE”
on the aluminum plate faces the batteries.
4.3 2010-A Distance Clip 60°
The 2010-A clip positions the fiberoptics end-piece relative to the
sample. The axis of the end-piece is positioned at a 60° angle
relative to the sample plane. Two different spacer rings may be
used to increase the distance between fiberoptics and sample.
In case of relative thick leaves, or when lichens and mosses are
examined, the sample may be placed below the hole of the
2010-A clip. Normal leaves are usually examined above this
Fig.
3: Distance Clip 60° 2010-A

Chapter 4 Components and Setup
12
hole. In the latter case, the leaf can be held between the folded
parts of the clip.
The distance between fiberoptics exit plane and sample has
considerable influence on signal amplitude and effective light in-
tensities (Fig. 4, page 12). With a 60° angle between sample
plane and fiberoptics, the distance between leaf surface and fiber
optics varies. Hence, the leaf surface is exposed to slightly het-
erogeneous light intensities when actinic light is applied via the
fiberoptics. A much more pronounced intensity gradient exists in-
side the leaf due to shading by the top chloroplast layers. In es-
sence, the measured signal will be dominated by that part of the
leaf which receives maximal intensity, as this also is most strong-
ly excited by the measuring light and emits most of the fluores-
cence which is received by the fiberoptics.
Fig.
4: Relationship between signal amplitude/light intensity and distance
between fiberoptics exit plane and sample

Components and Setup Chapter 4
13
4.4 Accessories
4.4.1 2035-B Leaf-Clip Holder
The Leaf-Clip Holder 2035-B must be connected to the LEAF
CLIP socket (Fig. 1, page 8) to record PAR, leaf temperature and
ambient humidity in parallel with chlorophyll fluorescence. In the
stand-alone mode, readings of environmental data are taken with
every saturation pulse analysis but these data can be continu-
ously recorded when the MINI-PAM-II is operated by the Win-
Control-3 software.
The Leaf-Clip Holder 2035-B is almost indispensable for field in-
vestigations, when ambient conditions may vary considerably. It
substitutes for the standard "Distance Clip" (2010-A) as a device
Fig.
5: 2035-B Leaf-Clip Holder

Chapter 4 Components and Setup
14
for defined positioning of the fiberoptics relative to the leaf plane.
Also, using the PAR sensor of the leaf clip, the internal PAR sen-
sor of the MINI-PAM-II can be readily calibrated.
In the 2035-B holder, the leaf is resting on a Perspex tube with
widened crest. The tube can be vertically adjusted to account for
different leaf thicknesses. The fiberoptics axis forms a 60° angle
with the leaf plane. Optionally, a 90° fiberoptics adapter (2030-
B90) is available for applications requiring homogenous illumina-
tion by actinic light applied via the fiberoptics. The distance be-
tween fiberoptics and leaf can be varied. For most applications
the minimal distance is recommended (maximal signal). Larger
distances can be defined by spacer rings. The illuminated leaf
area is limited by a steel ring with 10 mm ∅opening.
At the bottom of the Leaf-Clip Holder 2035-B, a tripod mounting
thread is provided. Mounting the device on a tripod (e. g. Com-
pact Tripod ST-2101A) facilitates long term measurements with
the same plant.
The handle of the Leaf-Clip Holder 2035-B features a red push-
button for remote control of the MINI-PAM-II. Pressing the button
triggers a saturation pulse and associated measurements of fluo-
rescence levels for “fluorescence quenching analysis”.
Micro-Quantum-Sensor
A micro quantum sensor is integrated into the Leaf-Clip Holder
2035-B to monitor the photosynthetic active radiation (PAR, be-
tween 400 and 700 nm) to which the sample is exposed. The mi-
cro-quantum-sensor measures light intensity in µmol quanta m-2
s-1. The µmol quanta m-2 s-1 is the unit of photon flux density.
Hence, the micro-quantum-sensor actually measures photosyn-
thetic photon flux density (PPFD).