MULTI-COLOR-PAM-II

Version:

Product

New

ST-Kinetics and Multi-Wavelength Chlorophyll Fluorometer

The new MULTI-COLOR-PAM-II is a compact instrument that houses two quite different techniques: the PAM technique and single turnover flash kinetics (STK).

MULTI-COLOR-PAM-II Highlights

Two instruments in one house: non-modulated flash analysis and multi-color PAM-modulated applications

A complete portfolio of fast measurements (time resolution down to 0.3 µs) to probe the PS II donor side and the presence of Car-triplets and multi-color PAM measurements to probe the PS II acceptor side, the electron transport chain and photosynthetic activity.

Mixing of both techniques: e.g. application of saturating 3 µs flashes, where the decay kinetics of the induced fluorescence signal can be monitored by PAM measuring light with a logarithmically declining measuring light frequency.

Overview of possible applications of the instrument

Pulse-amplitude Modulation (PAM)

Q_A-kinetics up to overall Photosynthesis activity

Single Turnover Flash Kinetics (STK)

Oxygen evolving complex up to PQ pool

Pulse-modulated fluorescence

Non-modulated fluorescence

Dark decay measurements

Pump-probe measurements

MT Pulses/continuous light

ST-flashes

Saturation pulse quenching analysis

Saturation flash quenching analysis

Regulatory fluorescence quenching (NPQ) and complementary PS II quantum yields

Short-lived fluorescence quenchers P680+ (DQ) and Car-triplets (TQ)

Dark-to-light induction curves

ST-kinetics (STK)

Two wavelength detection F_V(I)

Period-4 oscillations in FV(II)

Fast kinetics (50 µs to seconds) and Slow kinetics (seconds to hours)

Flash train-induced fluorescence changes

Sigma(II) (O-I₁ rise)

Sigma(II) (FRRF emulation)

Q_A– re-oxidation kinetics

S-state decay

PS II quantum yield

S-state distribution

Light saturation curves (LC)

Flash saturation curves

O-I₁-I₂-P/OJIP transients

Wide range of ST-intensities

PS II heterogeneity

ST-widths and ST-dark intervals

Mixed use of both techniques: e.g., applying an STK during an O-I₁-I₂-P transient

More choice is an important keyword for the new MULTI-COLOR-PAM-II: the new STK (single turnover kinetics) flashlamp and fast detector add reactions on the PS II donor side and Car triplets to the already broad portfolio. Fast switching between PAM and ST-Kinetics measurements gives synergy, offering a mixing of both techniques. The instrument is also flexible enough to emulate FRRF-type measurements or to produce Ramp-Method-type saturation pulses.

The instrument was designed for the measurement of suspensions, but people working with leaves were also not forgotten. The previous model had a leaf clip that can still be used for the new instrument. In addition, the STK-flashlamp also has a detector and can be used as a stand-alone application for leaf measurements.

Possible configurations:

The Multi-Color (PAM) configuration: the multi-color-emitter (MCP-II-E) and its detector (MCP-II-D1 or MCP-II-D2ST)
Two Wavelength Detection (PAM) configuration (suspensions): a second detector (either MCP-II-D1 or MCP-II-D2ST) is added to allow the detection of fluorescence at e.g. < 710 nm (mainly PS II) and > 700 nm (PS II + PS I), simultaneously. A leaf clip allowing two wavelength detection will be available in the near future.
Full configuration: STK-flashlamp (maximal flash intensities > 1.0 mol photons m^-2 s^-1) and combi-detector (MCP-II-EDST + MCP-II-D2ST) allowing detection of signals down to approx. 100 ns are added to the Multi-Color (PAM) configuration. This gives access to PS II donor side reactions.
Single Turnover Kinetics (STK) configuration for suspension measurements: STK-flashlamp (MCP-II-EDST) in combination with the combi-detector (MCP-II-D2ST).
Single Turnover Kinetics (STK) configuration for leaf measurements: The STK-flashlamp as stand-alone application: the STK-flashlamp contains a detector for fluorescence measurements from the sample surface, which makes it ideal for the measurement of leaves.

In the Multi-Color (PAM) configuration, the user has access to information about the wavelength dependent effective PS II antenna size: Sigma(II), reactions on the PS II acceptor side (QA^– re-oxidation) and electron flow along the electron transport chain (O-I₁-I₂-P/OJIP transients) but also to Saturation Pulse quenching analysis, dark-light induction and recovery curves, as well as light response curves, and this all for 5 different excitation and measuring light wavelengths (440, 480, 540, 590, 625 nm) plus white light that can be applied in any combination. This has the advantage that someone working, e.g., with diatoms can excite these organisms in the green and in the case of cyanobacteria either choose 625 nm to excite the phycobilisomes or 440 nm to excite the chlorophylls of the core antenna.

Not only the different wavelengths for fluorescence excitation and detection make the Multi-Color (PAM) configuration stand out. Other important features are:

The high sensitivity of the instrument, which allows measurements with optically thin samples, i.e., in the absence of intensity gradients of the various light qualities and largely avoiding wavelength-dependent fluorescence reabsorption – light gradient free.
Free choice of detection wavelengths using optical filters, e.g., enabling detection of fluorescence >700 nm, enriched in PS I fluorescence F(I), and < 710 nm, enriched in PS II fluorescence F(II), that can be measured simultaneously under equal conditions.
Characterization of the same state of a given sample by comparative PAM and flash kinetics (STK) measurements.
The almost complete freedom to create trigger and script files means that the experiments that can be designed are mainly limited by the creativity of the user.

New features of the MULTI-COLOR-PAM-II

The new elements of the MULTI-COLOR-PAM-II add whole new domains to the measurement portfolio of the instrument, making it even more a multi-function fluorometer.

High time resolution: The new STK-detectors provide a time resolution of 0.3 µs.
Time-resolved flash responses: the MULTI-COLOR-PAM-II is the first commercially available instrument with which the fluorescence yield during a saturating µs flash can be measured and carotenoid triplet quenching (TQ) and donor-side dependent quenching (DQ) can be differentiated.
Highly precise: For detailed analysis of the flash responses their timing is very precise and reproducible, and the form of the flashes approaches a rectangle, with the LEDs needing about 0.5 µs to reach full intensity. A special routine is provided for flash-profile correction.
Extremely intense flashes: Flash intensities of more than 1 000 000 µmol photons m^-2 s^-1 (440 nm) can be achieved with the EDST emitter-detector unit (yielding more than 1 excitation per µs).
Pump-probe: Double flash experiments with variable dark intervals ∆t (from 1 µs to 10 ms) allowing highly flexible relaxation measurements of various forms of quenching.
Period-4 oscillations: Flash frequencies of up to 100 Hz (10 ms time interval) can be used for flash trains to probe the S-states of the oxygen evolving complex.
Leaf measurements: Although the instrument was originally designed for suspension measurements, the EDST emitter-detector (STK-flashlamp) unit can also be used by itself to measure leaves. In addition, a configuration will be available soon in which two emitters are placed under a 45° angle relative to the multi-color emitter, which allows two wavelength detection of fluorescence emitted by leaves.
STK embedded in PAM recording: Thanks to fast switching [8-10 µs switching time] between fast non-modulated and PAM measurements an ST can be placed anywhere along a traditional – e.g. O-I₁-I₂-P or Slow Kinetics recording, with the resulting STK revealing details on the state of PS II at the moment of the flash. All this taken together makes the MULTI-COLOR-PAM-II a very all-round fluorometer allowing the user to probe and monitor PS II and the photosynthetic electron transport chain in many different ways.

Scientific Publications using Walz Devices

Source: Google Scholar.
Keywords: (Walz OR Waltz) Effeltrich.
Date: June 22, 2026.

Ʃ = 19642

Per Year

Source: Google Scholar.
Keywords: (Walz OR Waltz) Effeltrich.
Date: June 22, 2026.

Ʃ = 19642

Year

Selected Publications

Target of rapamycin is a crucial regulator of photosynthesis and nutrient metabolism partitioning in Nannochloropsis gaditana

https://doi.org/10.1186/s13068-025-02617-6

Biotechnology for Biofuels and Bioproducts 18: 21

Effect of overexcitation of photosystem II on chlorophyll fluorescence quenching parameters in Arabidopsis thaliana state transition mutants

Krieger-Liszkay A

Physiologia Plantarum 177: e70335

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NgLst8 coactivates TOR signaling to activate photosynthetic growth in Nannochloropsis gaditana

Zhang Z, Yang S, Li Y, Xie D, Chen G, Ren J, Zhu H, Zhou H

Microorganisms 12: 2574

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Redesign of the Chlamydomonas reinhardtii Q_B binding niche reveals photosynthesis works in the absence of a driving force for Q_A-Q_B electron transfer

Lambreva MD, Zobnina V, Antal TK, Peeva VN, Giardi MT, Bertalan I, Johanningmeier U, Virtanen O, Ray M, Mulo P, Polticelli F, Tyystjärvi E, Rea G

Physiologia Plantarum 176: e70008

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The role of protonation processes in the gradual formation of the light-adapted charge-separated state of photosystem II

Magyar M, Sipka G, Domonkos I, Chen X, Wang X, Han G, Shen J-R, Lambrev PH, Garab G

Frontiers in Photobiology 3: 1623224

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The photoprotective behavior of a motile benthic diatom as elucidated from the interplay between cell motility and physiological responses to a light microgradient using a novel experimental setup

Morelle J, Bastos A, Frankenbach S, Frommlet JC, Campbell DA, Lavaud J, Serôdio J

Microbial Ecology 87: 40

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Light-induced increase in the steady state chlorophyll fluorescence in cyanobacteria reflects induction of energy dissipation complementary to orange carotenoid protein-dependent thermal dissipation

Ogawa T, Takahashi H, Nishiyama Y, Hihara Y, Sonoike K

Photosynthesis Research 168: 38

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Influence of a novel organometallic Cu(II) complex on the photochemical activity of photosystem II in spinach

Shabanova M, Zharmukhamedov S, Allakhverdiev S

Transactions of the Institute of Molecular Biology & Biotechnologies 9: 47-51

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The effect of liquids activated by plasma generated with a microwave plasmatron and high-frequency glow discharge on cotton plant development

Shumeyko SA, Yanykin DV, Paskhin MO, Lukanin VI, Zakharov DA, Astashev ME, Pishchalnikov RY, Sarimov RM, Ashurov MK, Ashurov EM, Rashidova DK, Yakubov MM, Davydov AM, Gudkova VV, Danileyko YK, Dorokhov AS, Gudkov SV

Plants 14: 304

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Phosphorus starvation induces the synthesis of novel lipid class diacylglyceryl glucorunide and diacylglyceryl-N,N,N-trimethylhomoserine in two species of cold-adapted microalgae Rhaphidonema (Chlorophyta)

Suzuki H, Cuiné S, Légeret B, Wijffels RH, Hulatt CJ, Li-Beisson Y, Kiron V

The Plant Journal 121: e17227

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Nitrogen source type modulates heat stress response in coral symbiont (Cladocopium goreaui)

Huang Y, He J, Wang Y, Li L, Lin S

Applied and Environmental Microbiology 91: e00591-24

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Static and dynamic acclimation mechanisms to extreme light intensities in Hedera helix (Ivy) plants

Zer H, Ben-Ami AZ, Keren N

Physiologia Plantarum 177: e70217

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Relationship between harvesting efficiency and filament morphology in Arthrospira platensis Gomont

Kim G-H, Lee YJ, Kwon J-H

Microorganisms 13: 367

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Phytotoxic strains of Fusarium commune isolated from truffles.

Zvonarev A, Terentyev V, Zhelifinova V, Antipova T, Baskunov B, Avtukh A, Abashina T, Kachalkin A, Vainshtein M

Journal of Fungi 10: 465

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Metabolomics reveals the impact of overexpression of cytosolic fructose-1,6-biphosphatase on the photosynthesis and growth in Nannochloropsis gaditana.

Zhang Z, Li Y, Wen S, Yang S, Zhu H, Zhou H

International Journal of Molecular Sciences 25: 6800

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Synergistic CO₂ removal via enhanced olivine weathering and diatom growth in the ocean.

Zhang E, Li Y, Wang Y, Liu D, Cong Y, Liu J, Tang K, Nianzhi J, Zheng Q

Ocean-Land-Atmosphere Research 3: 0047

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A comprehensive study of light quality acclimation in Synechocystis sp. PCC 6803.

Zavřel T, Segečova A, Kovács L, Lukeš M, Novák Z, Pohland A-C, Szabó M, Somogyi B, Prášil O, Červený J, Bernát G

Plant & Cell Physiology 65: 1285-1297

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Effect of photoluminophore light-correcting coatings and bacterization by associative microorganisms on the growth and productivity of Brassica juncea L. Plants.

Zakharchenko NS, Rukavtsova EB, Yampolsky IV, Balakirev DO, Dyadishchev IV, Ponomarenko SA, Luponosov YN, Filonov AE, Mikhailov PA, Zvonarev AN, Akhmetov LI, Terentyev VV, Khudyakova AY, Zalomova LV, Tarlachkov SV, Aripovsky AV, Puntus IF, Khramov RN

Microbiology Research 15: 1957-1972

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Engineering RNA polymerase to construct biotechnological host strains of cyanobacteria.

Turunen O, Saleem T, Kurkela J, Kallio P, Tyystjärvi

Physiologia Plantarum 176: e14263

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Ocean deoxygenation dampens resistance of diatoms to ocean acidification in darkness.

Sun J-Z, Zhang D, Yi X, Beardall J, Gao K

Frontiers in Marine Science 11: 1387552

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Shedding light on blue-green photosynthesis: A wavelength-dependent mathematical model of photosynthesis in Synechocystis sp. PCC 6803.

Pfennig T, Kullmann E, Zavřel T, Nakielski A, Ebenhöh O, Červený J, Bernat G, Matuszyńska AB

PLOS Computational Biology 20: e1012445

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Low-CO₂-inducible bestrophins outside the pyrenoid sustain high photosynthetic efficacy in diatoms.

Nigishi M, Shimakawa G, Yamagishi K, Amano R, Ito S, Tsuji Y, Nagasato C, Matsuda Y

Plant Physiology 195: 1432-1445

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NgAP2a targets KCS gene to promote lipid accumulation in Nannochloropsis gaditana.

Lin Y, Li Y, Wu X, Xu W, Zhang Z, Zhu H, Zhou H

International Journal of Molecular Sciences 25: 10305

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Thermal responses of Tetradesmus obliquus for industrial outdoor cultivation.

Kato H, Suzuki H, Wijffels RH, Schulze PSC, Hulatt CJ

Bioresource Technology Reports 27: 101909

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Zeaxanthin epoxidase 3 knockout mutants of the model diatom Phaeodactylum tricornutum enable commercial production of the bioactive carotenoid diatoxanthin.

Graesholt C, Brembu T, Volpe C, Bartosova Z, Serif M, Winge P, Nymark M

Marine Drugs 22: 185

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Phylogenetic profiling analysis of the phycobilisome revealed a novel state-transition regulator gene in Synechocystis sp. PCC 6803.

Fukunaga T, Ogawa T, Iwasaki W, Sonoike K

Plant & Cell Physiology 65: 1450-1460

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Marine heatwave-driven mass mortality and microbial community reorganization in an ecologically important temperate sponge.

Bell JJ, Micaroni V, Strano F, Ryan KG, Mitchell K, Mitchell P, Wilkinson S, Thomas T, Bachtiar R, Smith RO

Global Change Biology 30: e17417

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Light-dependent methane production by a coccolithophorid may counteract its photosynthetic contribution to carbon dioxide sequestration

Rao Y, Gao G, Berman-Frank I, Bizic M, Gao K

Communications Earth & Environment 5: 695

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Flash-kinetics as a complementary analytical tool in PAM fluorimetry

Christof Klughammer, Friedemann Schlosser, Ulrich Schreiber

Photosynthesis Research 161: 151-176

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Probing the influence of novel organometallic copper(II) complexes on spinach PS II photochemistry using OJIP fluorescence transient measurements.

Zharmukhamedov SK, Shabanova MS, Huseynova IM, Karacan MS, Karacan N, Akar H, Kreslavski VD, Alharby HF, Bruce BD, Allakhverdiev SI

Biomolecules 13: 1058

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Photo-physiological acclimation in Synechocystis sp. PCC 6803 provides insight into growth limitation in underwater spectra.

Zavřel T, Segečová A, Kovács L, Lukeš M, Novák Z, Szabó M, Somogyi B, Prášil O, Červený J, Bernát G

bioRxiv

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Salinity-induced changes in plastoquinone pool redox state in halophytic Mesembryanthemum crystallinum L.

Pilarska M, Niewiadomska E, Kruk J

Scientific Reports 13: 11160

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Loss of CpFTSY reduces photosynthetic performance and affects insertion of PsaC of PS I in diatoms.

Nymark M, Finazzi G, Volpe C, Serif M, de Mirandi Fonseca D, Sharma A, Sanchez N, Sharma AK, Ashcroft F, Kissen R, Winge P, Bones AM

Plant & Cell Physiology 64: 583-603

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Photoinhibition of the picophytoplankter Synechococcus is exacerbated by ocean acidification.

Li H, Beardall J, Gao K

Water 15: 1228

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Micractinium lacustre and M. thermotolerans spp. nov. (Trebouxiophyceae, Chlorophyta): taxonomy, temperature-dependent growth, photosynthetic characteristics and fatty acid composition.

Krivina E, Sinetova M, Savchenko T, Degtyaryov E, Tebina E, Temraleeva A

Algal Research 71: 103042

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Light map optimization via direct chlorophyll fluorescence imaging in algal photobioreactors.

Kofler JR, Labeeuw L, Bates H, Zavafer A, Ralph PJ

Algal Research 71: 103022

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A possible relationship between the effect of factors on photoactivation of photosystem II depleted of functional Mn and cytochrome b₅₅₉.

Khorobrykh A

Biochimica et Biophysica Acta – Bioenergetics 1864: 148997

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Phytoplankton tune local pH to actively modulate circadian swimming behavior.

Ghoshal A, Dhar J, Grossart H-P, Sengupta A

bioRxiv

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The slow-phase of chlorophyll fluorescence induction curve reflects the electron transport rates of photosystem II in vivo in Chlorella vulgaris.

Bates H, Zavafer A, Szabo M, Ralph PJ

Journal of Applied Phycology 35: 109-116

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Light-induced changes of far-red excited chlorophyll fluorescence: further evidence for variable fluorescence of photosystem I in vivo.

Schreiber U

Photosynthesis Research 155: 247-270

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Active reconfiguration of cytoplasmic lipid droplets governs migration of nutrient-limited phytoplankton.

Sengupta A, Dhar J, Danza F, Ghoshal A, Müller S, Kakavand N

Science Advances 8: eabn6005

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Reduced salinity exacerbates the viral infection on the coccolithophorid Emiliania huxleyi at elevated pCO₂.

Fu Q, Gao K

Frontiers in Marine Science 9: 1091476

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A novel algicidal bacterium, Microbulbifer sp. YX04, triggered oxidative damage and autophagic cell death in Phaeocystis globose, which causes harmful algal blooms.

Zhu X, Chen S, Luo G, Zheng W, Tian Y, Lei X, Yao L, Wu C, Xu H

Microbiology Spectrum 10: e00934

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Effects of three graphene-based materials on the growth and photosynthesis of Brassica napus L.

Xiao X, Wang X, Liu L, Chen C, Sha A, Li J

Ecotoxicology and Environmental Safety 234: 113383

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Perspective design of algae photobioreactor for greenhouses – a comparative study.

Sukačová K, Lošák P, Brummer V, Máša V, Vícha D, Zavřel T

Energies 14: 1338

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An insight into the bicarbonate effects in photosystem II through the prism of the JIP test.

Shitov AV

Photochem 2: 779-797

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Hysteresis light curves: a protocol for characterizing the time dependence of the light response of photosynthesis.

Serôdio J, Moreira D, Bastos A, Cardoso V, Frommlet J, Frankenbach S

Photosynthesis Research 154: 57-74

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Nitrogen limitation enhanced calcification and sinking rate in the Coccolithophorid Gephyrocapsa oceanica along with its growth being reduced.

Jiang X, Li H, Tong S, Gao K

Frontiers in Marine Science 9: 834358

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Differences in susceptibility to photoinhibition do not determine growth rate under moderate light in batch or turbidostat – a study with five green algae.

Mattila H, Valev D, Mishra KB, Havurinne V, Virtanen O, Antinluoma M, Tyystjärvi E

Photosynthetica 60: 10-20

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Metabolic alterations in alga Chlamydomonas reinhardtii exposed to nTiO₂ materials.

Liu W, Li M, Li W, Keller AA, Slaveykova VI

Environmental Science Nano 9: 2922

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Heterologous lactate synthesis in Synechocystis sp. Strain PCC 6803 causes a growth condition-dependent carbon sink effect.

Grund M, Jakob T, Toepel J, Schmid A, Wilhelm C, Bühler B

Applied and Environmental Microbiology 88: 0063

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Monitoring fitness and productivity in cyanobacteria batch cultures.

Zavřel T, Schoffman H, Lukeš M, Fedorko J, Keren N, Červený J

Algal Research 56: 102328

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Transcriptomic analysis of Chlorella sp. HS2 suggests the overflow of acetyl-CoA and NADPH cofactor induces high lipid accumulation and halotolerance

Yun J-H, Pierrelée M, Cho D-H, Kim U, Heo J, Choi D-Y, Lee YJ, Lee B, Kim HR, Habermann B, Chang YK, Kim H-S

Food and Energy Security 10: e267

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Harmful algal bloom-forming dinoflagellate Prorocentrum donghaiense inhibits the growth and photosynthesis of seaweed Sargassum fusiformis embryos.

Wang C, Wang M, Chen B, Qin W, Lin L, Dai C, Yu H, Li R, Zhao M, Ma Z

Journal of Oceanology and Limnology 39: 2237-2251

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Marine Synechococcus picocyanobacteria: light utilization across latitudes.

Six C, Ratin M, Marie D, Corre E

Proceedings of the National Academy of Sciences USA 118: e2111300118

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Evidence for variable chlorophyll fluorescence of photosystem I in vivo.

Schreiber U, Klughammer C

Photosynthesis Research 149: 213-231

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Elevated pCO₂ enhances under light but reduces in darkness the growth rate of a diatom, with implications for the fate of phytoplankton below the photic zone.

Qu L, Beardall J, Jiang X, Gao K

Limnology and Oceanography 66: 3630-3642

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Underwater light climate and wavelength dependence of microalgae photosynthetic parameters in a temperate sea.

Michel-Rodriguez M, Lefebvre S, Crouvoisier M, Mériaux X, Lizon F

PeerJ 9: e12101

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Interactions between ultraviolet B radiation, warming, and changing nitrogen source may reduce the accumulation of toxic pseudo-nitzschia multiseries biomass in future coastal oceans.

Kelly KJ, Fu F-X, Jiang X, Li H, Xu D, Yang N, DeMers MA, Kling JD, Gao K, Ye N, Hutchins DA

Frontiers in Marine Science 8: 664302

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Validation of parameters and protocols derived from chlorophyll a fluorescence commonly utilised in marine ecophysiological studies.

González-Guerrero LA, Vásquez-Elizondo RM, López-Londoño, Hernán G, Iglesias-Prieto R, Enríquez S

Functional Plant Biology 49: 517-532

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Photomorphogenesis in the picocyanobacterium Cyanobium gracile includes increased phycobilisome abundance under blue light, phycobilisome decoupling under near far-red light, and wavelength-specific photoprotective strategies.

Bernát G, Zavřel T, Kotabová E, Kovács L, Steinbach G, Vörös L, Prášil O, Somogyi B, Tóth VR

Frontiers in Plant Science 12: 612302

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Interactive effects of elevated CO₂ concentration and light on the picophytoplankton Synechococcus.

Bao N, Gao K

Frontiers in Marine Science 8: 634189

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Light availability modulates the effects of warming in a marine N₂ fixer.

Yi X, Fu F-X, Hutchins DA, Gao K

Biogeosciences 17: 1169-1180

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Lower salinity leads to improved physiological performance in the Coccolithophorid Emiliania huxleyi, which partly ameliorates the effects of ocean acidification.

Xu J, Sun J, Beardall J, Gao K

Frontiers in Marine Science 7: 704

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Polychromatic Fourier-PAM fluorometry and hyperspectral analysis of chlorophyll fluorescence from Phaseolus vulgaris leaves: effects of green light.

Lysenko V, Guo Y, Koslapov A, Usova E, Varduny T, Krasnov V

Information Processing in Agriculture 7: 204-211

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Synoptic spatio-temporal variability of the photosynthetic productivity of microsphytobenthos and phytoplankton in a tidal estuary.

Frankenbach S, Ezequiel J, Plecha S, Goessling JW, Vaz L, Kühl M, Dias JM, Vaz N, Serôdio J

Frontiers in Marine Science 7: 170

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Light stress in green and red Plankothrix strains: the orange carotenoid protein and its related photoprotective mechanism.

Djediat C, Feilke K, Brochard A, Caramelle L, Tiam SK, Sétif P, Gauvrit T, Yéprémian C, Wilson A, Talbot L, Marie B, Kirilovsky D, Bernard C

Biochimica et Biophysica Acta 1861: 148037

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Astaxanthin biosynthesis in transgenic Dunaliella salina (Chlorophyceae) enhanced tolerance to high irradiation stress.

Chen Y, Bi C, Zhang J, Hou H, Gong Z

South African Journal of Botany 133: 132-138

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Unresolved quenching mechanisms of chlorophyll fluorescence may invalidate MT saturating pulse analyses of photosynthetic electron transfer in microalgae.

Havurinne V, Mattila H, Antinluoma M, Tyystjärvi E

Physiologia Plantarum 166: 365-379

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Rapidly reversible chlorophyll fluorescence quenching induced by pulses of supersaturating light in vivo.

Schreiber U, Klughammer C, Schansker G

Photosynthesis Research 142: 35-50

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Unique photosynthetic electron transport tuning and excitation distribution in heterokont algae.

Røkke GB, Melø TB, Mühlroth A, Vadstein O, Bones AM, Hohmann-Marriott MF

PLOS ONE 14: e0209920

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Isolation and characterization of novel Chlorella species with cold resistance and high lipid accumulation for biodiesel production.

Koh HG, Kang NK, Kim EK, Suh WI, Park W-K, Lee B, Chang YK

Journal of Microbial Biotechnology 29: 952-961

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Effect of carbon limitation on photosynthetic electron transport in Nannochloropsis oculata.

Zavřel T, Szabo M, Tamburic B, Evenhuis C, Kuzhiumparambil U, Literáková P, Larkum AWD, Raven JA, Červeny, Ralph PJ

Journal of Photochemistry and Photobiology B 181: 31-43

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Structure-based optics of centric diatom frustules: modulation of the in vivo light field for efficient diatom photosynthesis.

Goessling JW, Su Y, Cartaxana P, Maibohm C, Rickelt LF, Trampe ECL, Walby SL, Wangpraseurt D, Wu X, Ellegaard M, Kühl M

New Phytologist 219: 122-134

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Truncated light-harvesting chlorophyll antenna size in Chlorella vulgaris improves biomass productivity.

Shin W-S, Lee B, Jeong B-r, Chang YK, Kwon J-H

Journal of Applied Phycology 28: 3193-3202

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A radiative transfer modeling approach for accurate interpretation of PAM fluorometry experiments in suspended algal cultures.

Murphy TE, Prufert-Bebout LE, Bebout BM

Biotechnology Progress 32: 1601-1608

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Photoregulation in a kleptochloroplastidic dinoflagellate, Dinophysis acuta.

Hansen PJ, Ojamäe K, Berge T, Trampe ECL, Nielsen LT, Lips I, Kühl M

Frontiers in Microbiology 7: 785

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Photo-protection in the centric diatom Coscinodiscus granii is not controlled by chloroplast high-light avoidance movement.

Goessling JW, Cartaxana P, Kühl M

Frontiers in Marine Science 2: 115

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Apparent PS II absorption cross-section and estimation of mean PAR in optically thin and dense suspensions of Chlorella.

Klughammer C, Schreiber U

Photosynthesis Research 123: 77-92

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Spectral Effects on Symbiodinium photobiology studied with a programmable light engine.

Wangpraseurt D, Tamburic B, Szabó M, Suggett D, Ralph PJ, Kühl M

PLOS ONE 9: e112809

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Effective light absorption and absolute electron transport rates in the coral Pocillopora damicornis.

Szabó M, Wangpraseurt D, Tamburic B, Larkum AWD, Schreiber U, Suggett DJ, Kühl M, Ralph PJ

Plant Physiology and Biochemistry 83: 159-167

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Photosynthetic acclimation of Nannochloropsis oculata investigated by multi-wavelength chlorophyll fluorescence analysis.

Szabó M, Parker K, Guruprasad S, Kuzhiumparambil U, Lilley RMcC., Tamburic B, Schliep M, Larkum AWD, Schreiber U, Raven JA, Ralph PJ

Bioresource Technology 167: 521-529

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Wavelength-dependent photodamage to Chlorella investigated with a new type of multi-color PAM chlorophyll fluorometer.

Schreiber U, Klughammer C

Photosynthesis Research 114: 165-177

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Assessment of wavelength-dependent parameters of photosynthetic electron transport with a new type of multi-color PAM chlorophyll fluorometer.

Schreiber U, Klughammer C, Kolbowski J

Photosynthesis Research 113: 127-144

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Unique properties vs. common themes: the atypical cyanobacterium Gloeobacter violaceus PCC 7421 is capable of state transitions and blue-light induced fluorescence quenching.

Bernát G, Schreiber U, Sendtko E, Stadnichuk IN, Rexroth S, Rögner M, Koenig F

Plant and Cell Physiology 53: 528-542

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MULTI-COLOR-PAM-II

General design

ARM microcontroller (480 MHz), Fast Kinetics up to 128,000 points with 14 bit resolution, ST Kinetics up to 15,000 points with 14 bit resolution, unlimited storage for Slow Kinetics

Sockets

4 sockets for measuring light and actinic light of MCP-II-E Multi-Color Emitter Head and measuring light/auxiliary detector and ST supply MCP-II-EDST, 2 sockets for signal detection by MCP-II-D1 and MCP-II-D2DST Detector Heads, charge socket or Battery Charger MINI-PAM/L, output socket for PHYTO-MS Miniature Magnetic Stirrer, 4 BNC sockets for 5 V trigger in and trigger out signals and ST trigger in and trigger out signals, input socket for US-SQS/WB Spherical Micro Quantum Sensor or US-MQS/WB Mini Quantum Sensor, input socket for auxiliary devices, USB socket

Communication

USB 2.0, USB 3 compatible

User interface

Windows computer with PamWin-4 software

Power supply

standard: NO sealed lead-acid battery; Battery Charger MINI-PAM/L (100 to 240 V AC)

Dimensions

31 cm x 16 cm x 33.5 cm (W x H x D)

Power consumption

6 W

Weight

3.64 kg

Operating temperature

-5 to +40 °C

Chip-on-board multi-wavelength measuring light LED emitter

400, 440, 480, 540, 590, 625 nm for pulse-modulated measuring light; 20 intensity settings and 14 pulse frequency settings

Chip-on-board multi-wavelength actinic LED array

440, 480, 540, 590, 625 and 420-640 nm (white) for continuous actinic illumination, max. 5000 µmol photons m-2 s-1; saturating single turnover flashes, max. 200000 µmol photons m-2 s-1; multiple turnover pulses, max. 12000 µmol m-2 s-1 PAR, adjustable between 1 and 800 ms

Far-Red LED

peak wavelength 730 nm

Dimensions

10.5 cm x 5.5 cm x 7 cm (L x W x H)

Weight

500 g (incl. cables, 1 m long)

Signal detection

PIN photodiode with special pulse preamplifier for measuring fluorescence changes with maximum time resolution of 10 µs

Filter box

for up to 14 mm filter thickness

Standard detector filter

long-pass filter >650 nm (3 mm RG 665) plus short-pass filter SP 710

Dimensions

6.9 cm x 9.8 cm x 6.4 cm (L x W x H)

Weight

355 g (incl. cables, 1 m long)

Signal generation and detection

High quality 450 nm ST-flashes of variable duration and intensity up to 1 mol photons m-2 s-1; Actinic light (450 nm); Far Red light (740 nm); STK-detector for surface detection of direct fluorescence with sub-µs time resolution

Dimensions

15.4 cm x 5.2 cm x 7 cm

Weight

570 g (incl. cables, 1.2 m long)

STK & PAM-Detector (switchable)

includes: PAM-detector equivalent to MCP-II-D1 and STK-detector for non-modulated fluorescence with sub-µs time resolution

Filter box

for up to 14 mm filter thickness

Standard detector filter

long-pass filter >650 nm (3 mm RG 665) plus short-pass filter SP 710

Dimensions

6.9 cm x 9.8 cm x 6.4 cm (L x W x H)

Weight

550 g (incl. cables, 1.2 m long)

Input

90 to 264 V AC, 47 to 63 Hz

Output

19 V DC, 3.7 A

Operating temperature

0 to 40 °C

Dimensions

15 cm x 6 cm x 3 cm (L x W x H)

Weight

300 g

Software

PamWin-4 System Control and Data Acquisition Program for operation of the instrument via PC or laptop, data acquisition and analysis

Fluorescence analysis

Fitting routine for the fast fluorescence O-I1 to determine the functional absorption cross-section of the PS II antennae (Sigma) needed for the determination of PS II-specific electron transport rates

Fitting routine for exponential decay (e.g. QA– reoxidation after a single turnover flash) or rise of a signal with up to three exponentials

Choice of two fitting routines for light curves (determination of cardinal points α, Ik and ETRmax)

Measured

Ft, F0, FM, F, F0' (also calculated), FM'. Fast polyphasic rise and decay kinetics (time resolution up to 10 µs). PAR using Spherical Micro Quantum Sensor US-SQS/WB or Mini Quantum Sensor US-MQS/WB

Calculated

F0' (also measured), Fv/Fm and Y(II) (maximum and effective photochemical yield of PS II, respectively), qL, qP, qN, NPQ, Y(NPQ), Y(NO) and ETR (electron transport rate), C/F0 (constant fraction of F₀ that is assumed to originate from PS I)

Measured

Ft, F₀, F_M, F, F₀' (also calculated), F_M'. Fast polyphasic rise and decay kinetics (time resolution up to 10 μs). PAR using Spherical Micro Quantum Sensor US-SQS/WB or Mini Quantum Sensor US-MQS/WB.

Calculated

F₀' (also measured), F_V/F_M and Y(II) (maximum and effective photochemical yield of PS II, respectively), qL, qP, qN, NPQ, Y(NPQ), Y(NO) and ETR (electron transport rate), C/F₀ (constant fraction of F₀ not constituting PS II chlorophyll fluorescence)

Measured

Ft, F0, FM, F, F0’ (also calculated), FM’. Time-resolved Saturation Pulse: Fast polyphasic rise and decay kinetics (time resolution up to 10 µs). PAR using Spherical Micro Quantum Sensor US-SQS/WB or Mini Quantum Sensor US-MQS/WB

Calculated

F0’ (also measured), FV/FM and Y(II) (maximum and effective photochemical yield of PS II, respectively), qL, qP, qN, NPQ, Y(NPQ), Y(NO) and ETR (electron transport rate), C/F0 (constant fraction of F0 not constituting PS II chlorophyll fluorescence)

Operating System

Microsoft Windows 10 (32 and 64 bits) and 11; computers that can run these operating systems, can also work with PamWin-4.

Design

Aluminum box with custom foam packing

Dimensions

60 cm x 40 cm x 34 cm (L x W x H);
60 liter

Weight

5 kg

Dimensions

Base plate, 40 cm x 30 cm

Height

73.5 cm, diameter 1.5 cm

Weight

2.8 kg

Accessories

Design

Black-anodized aluminum body with central 10 x 10 mm glass cuvette; for attachment of Measuring Heads and Miniature Magnetic Stirrer PHYTO-MS; additional ports for attachment of two additional measuring heads

Weight

750 g

Temperature Control Block ED-101US/T

Sectioned block with central 10 x 10 mm opening to be mounted on top of the ED-101US/MD unit; to be connected to external flow-through water bath (not included), weight: 250 g

Miniature Magnetic Stirrer PHYTO-MS

Based on a device manufactures by h+p (type Variomag-Mini); featuring adapter to be mounted in the bottom port of the Optical Unit ED-101US/MD; powered and controlled by the Power-and-Control-Unit

Spherical Micro Quantum Sensor US-SQS/WB

3.7 mm Ø diffusing sphere coupled to integrated PAR sensor via 2 mm diameter fiber; compact amplifier unit and special holder for mounting on Optical Unit ED-101US/MD; to be connected tot the Power-and-Control Unit

Temperature Control Block ED-101US/T

Sectioned block with central 10 x 10 mm opening to be mounted on top of the ED-101US/MD unit; to be connected to external flow-through water bath (not included), weight 250 g

Miniature Magnetic Stirrer PHYTO-MS

Based on device manufactured by h+p (type Variomag-Mini); featuring adapter to be mounted in bottom port of the Optical Unit ED-101US/MD; powered and controlled by the Power-and-Control-Unit DKN-C

Spherical Micro Quantum Sensor US-SQS/WB

3.7 mm ø diffusing sphere coupled to integrated PAR sensor via 2 mm diameter fiber; compact amplifier unit and special holder for mounting on Optical Unit ED-101US/MD; to be connected to the Power-and-Control Unit DKN-C

Design

Magnetic stirrer driven by a rotating magnetic field; the PHYTO-MS is connected to Power-and-Control Unit PHYTO-II-C; a special adapter plug allows the insertion in the bottom port of the Optical Unit ED-101US/MP

Weight

16 g

Display

Three line LCD display

Control range

0 °C to 50 °C at 0.1 K steps

Operating voltage

11 V – 14 V DC

Maximum Peltier current

1 A

Size

105 mm x 90 mm x 130 mm (W x H x D)

Weight

0.57 kg

Power-and-Control Unit US-T/DR

Display

Three line LCD display

Control range

0 °C to 50 °C at 0.1 K steps

Operating voltage

11 V - 14 V DC

Maximum Peltier current

1 A

Size

105 mm x 90 mm x 130 mm (W x H x D)

Weight

0.57 kg

Peltier Heat-Transfer Head US-T/DS

Achievable temperatures

12 K below ambient temperature, 15 K above ambient temperature (Quartz cuvette placed in Optical Unit for Suspensions ED-101US/MD with 1.5 mL water and stirrer PHYTO-MS on)

Size

⌀ 55 mm, 110 mm height

Cable length

130 cm

Weight

0.29 kg (including cable)

AC Adapter

Input

100 V - 240 V AC 1.5 A 50-60 Hz

Output

12 V DC 5.5 A

Size

130 mm x 56 mm x 30 mm (L x W x H)

Weight

0.50 kg (including cable)

PamWin-4 Software

General Features and Graphical User Interface

PamWin-4, the software controlling both the MULTI-COLOR-PAM(-II) and the PAM-2500, was developed out of PamWin-3 and can handle all the new features of the MULTI-COLOR-PAM-II. The software runs on PCs with operating systems Windows 10 (32 and 64 bit) and 11.

The software consists of 3 parts: SP analysis, Fast Acquisition for PAM-applications (first 3 Tabs) and Single Turnover Kinetics (STK) for non-modulated flash analyses (last 2 Tabs).

The broad portfolio of applications also means a considerable number of sections on Tabs where particular applications can be defined. On the new ST Settings Tab single, double or multiple flash experiments can be defined (first example below), but also flash trains for the determination of period-4 oscillations (with or without pre-flashes) as shown in the second example below.

Simple menus to define flash experiments and flash trains with or without pre-flashes.

A simple menu allows the user to define a single or double (or multiple) flash experiment. The latter is particularly important for configuring pump-probe measurements

With the Pulse Series menu, a flash train can be defined. Setting the Target No. to 14, a flash train of 14 flashes, in this case 200 ms apart will be applied.
Giving two pre-flashes, here 100 ms apart, the S3 state can be induced. By varying the delay time between pre-flashes and flash train (here 1 s), the S3 state decay can be monitored. — With the Pulse Series menu, a flash train can be defined. Setting the Target No. to 14, a flash train of 14 flashes, in this case 200 ms apart will be applied. Giving two pre-flashes, here 100 ms apart, the S₃ state can be induced. By varying the delay time between pre-flashes and flash train (here 1 s), the S₃ state decay can be monitored.

Tools for the analysis of period-4 oscillations are part of the software (see Applications).

For the Fast Acquisition data, the software offers two types of support. On the one hand measurement protocols in the form of trigger files and scripts, and on the other hand fit routines for the analysis of O-I1-transients to determine the antenna cross section Sigma(II), the exponential fluorescence decay following a single turnover flash, or following a period of illumination.

The script and trigger files that come with the software can also be used as examples on the basis of which other script and trigger files can be developed.

Trigger file for a 300 ms polyphasic rise with a single turnover flash after 1 ms.

In the case of a Trig-run the experimental protocol defined by a trigger file is executed on the basis of the settings defined in the General Settings page.

In the case of a script file, one or more trigger files are imbedded in a timeline defining as well, the timing of the different parts of the experiment. In a script the user can also define the intensities and/or wavelengths of the different light sources used; settings, which can be changed as a function of the timeline of the script. Here, an example of a script file for a Sigma(II) determination, allowing for example the measurement of the five different excitation wavelengths in one go.

Suspension and leaf configurations

Scheme of a complete configuration for suspension measurements (a) and stand-alone application of the STK flashlamp unit for leaf measurements (b) (see Klughammer, Schlosser and Schreiber (Photosynth Res 2024: https://doi.org/10.1007/s11120-024-01101-w) for a full description).

In the schematic version of the stand-alone application (see above), the leaf is placed in a gas flow chamber.

For leaf fluorescence measurements with the Multi-Color Emitter a new leaf clip (MCP-II-BK) is available. The Multi-Color Emitter is placed in the center and the two detectors are placed under a 45° angle relative to the emitter as shown below. The leaf is kept in place with magnets located in the clip and the lid that is placed on the other side of the leaf. The old MCP-BK leaf clip is also compatible with the MULTI-COLOR-PAM-II.

Application

Example experiments from the three application domains

The software splits the applications of the MULTI-COLOR-PAM-II in 3 parts:

Photosynthetic Activity related applications like induction (+ recovery) curves and light curves, but also manual measurements.
Script-based experiments like O-I₁-I₂-P/OJIP transients, re-oxidation kinetics of Q_A^- following a single turnover flash, Sigma(II) determinations.
Flash-based experiments like period-4 oscillations, Car-triplet decay and induction kinetics, P680⁺
The first two parts represent PAM-applications, and the last part represents Single Turnover Kinetics (STK) applications.

Example of a Light Curve/Quenching Analysis

The first figure gives an example of a light curve recording of the complementary PS II quantum yields, Y(II)+Y(NPQ)+Y(NO)=1. where the light intensity was first increased and then decreased again. Due to memory effects the kinetics induced by the decreasing light intensities may differ from those of the increasing light intensities. This phenomenon is called hysteresis. The here observed full reversibility of light-induced lowering of Y(II) and increase of Y(NPQ) is characteristic for physiologically healthy samples.

Examples of Fast Kinetics (PAM) measurements

Two wavelength measurements of O-I₁-I₂-P transients

An example of a fast PAM-application is the simultaneous measurement of O-I₁-I₂-P transients in two different wavelength domains: < 710 nm and > 700 nm, where the fluorescence measured at wavelengths < 710 nm is mainly PS II fluorescence and the >700 nm fluorescence is a potential mix of PS II and PS I fluorescence. First a measurement of a dilute suspension of Chlorella cells (440 nm ML and MT) taken from Klughammer et al. (2024).

The O-I₁ rise (curves normalized to I₁) is the same for both wavelengths. A difference is observed between I₂ and P: the I₂-P rise is more pronounced at F>700 nm compared with F<710 nm.

Below, a barley leaf is measured using 440 nm measuring and actinic light using the two-wavelength configuration for leaves (see Configurations).

The two measurements were again normalized to I₁ (all Q_A reduced). Again, the I₂-P rise is more pronounced at F>700 nm compared with F<710 nm. The somewhat slower O-I₁ rise kinetics reflect the fact that self-absorption at F<710 nm is higher than at F>700 nm and, therefore, F>700 nm originates from relatively deeper layers in the leaf, where the effective actinic light intensity is lower.

Sigma(II) determination

The parameter Sigma(II) reflects the effective cross section of the PS II antenna. The Sigma(II) determination (and its wavelength dependence) is another fast PAM-application. There are three criteria on the basis of which one can judge if the O-I₁ fit used for the Sigma(II) determination was good: 1. The fit should describe the fluorescence rise well, 2. The obtained fit parameters should be physiologically relevant and 3. The obtained Sigma(II) values should be independent of the light intensity. Here, the Sigma(II) values were observed to increase with the age of the cultures used.

For this experiment, the connectivity parameter J was fixed to 1.2, the value obtained by Anne and Pierre Joliot in 1964. The dataset shows near perfect fits, reasonable parameter values and in essence an independence of the light intensity at high light intensities, yielding well-defined O-I1 kinetics. — For this experiment, the connectivity parameter J was fixed to 1.2, the value obtained by Anne and Pierre Joliot in 1964. The dataset shows near perfect fits, reasonable parameter values and in essence an independence of the light intensity at high light intensities, yielding well-defined O-I₁ kinetics.

STK, single turnover flash kinetics, applications

Car-triplet decay

The next figure shows a set of measurements on the basis of which the Car-Triplet decay kinetics can be determined. The dataset illustrates the precision of the timing of the flashes and the ability of the flash lamp to give two equally strong flashes 1 µs apart.

Flash length and fluorescence induction

Another example is a double flash experiment in which the length of the first flash was varied and the second flash is given 40 µs after the first flash.

Flash trains and flash patterns can tell us something about the S-states, the redox states of the manganese cluster on the donor side. They can also tell us something about the effects of different intensities of far red.

Period-4 oscillations

Period-4 oscillations in either F₀, F_M or F_V level can be derived automatically by the software (figure taken from Klughammer et al. 2024).

In coffee leaves, FR1 illumination already leads to a strong damping of the period-4 oscillations in the variable fluorescence. In such cases, the MULTI-COLOR-PAM-II allows a further reduction of the effective FR-intensity to 10% of FR1, which for this coffee leaf strongly reduced the effect on the S-states.

Combining STK and PAM measurements

In the next example it is shown how a mixing of STKs and PAM measuring light allows the combination of the precise, intense and short STKs and PAM measuring light allowing the monitoring of fluorescence decay in darkness.

3 µs STK flashes applied to a dilute Chlorella sample inhibited by DCMU (blue) or uninhibited (red) embedded in a PAM measurement. The ML-frequency declined logarithmically from 100 kHz to 10 kHz starting 100 µs after the flash (figure taken from Klughammer et al. 2024).

Usage

Water

Laboratory

Photosynthetic Light Reactions

Suspensions (High Conc.)

Algae

Highlights

PAM fluorescence and highly time-resolved continuous fluorescence
Advanced Photosystem II analysis

Downloads

Software: PamWin 4 04c Software
Manuals & Documentation: MULTI-COLOR-PAM-II Manual
PAM Application Notes (PAN): PAN (2011) 1: 1-19

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