Johan Sebastiaan Ploem

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Johan Sebastiaan Ploem
Born (1927-08-25) August 25, 1927 (age 89)
Sumatra, Indonesia
Nationality Dutch
Fields Advanced Microscopy; Molecular Diagnostics; Digital Painting
Institutions University of Miami; University of Amsterdam; University of Leiden
Notable awards Fellow of the Papanicolaou Cancer Research Institute
C. E. Alken Foundation award Ernst Abbe medal and award of the New York Microscopical Society Erica Wachtel medal from the British Society for Clinical Cytology The first honorary member of the Society for Analytical Cytology Honorary fellow of the Royal Microscopical Society Honorary member of the New York Microscopical Society


Johan Sebastiaan Ploem (born 25 August 1927, Sawahlunto) is a Dutch microscopist and digital artist, who made significant contribution to the field of fluorescence microscopy.

Personal life[edit]

Ploem was born on Sumatra, then part of the Dutch East Indies. He started painting as a small boy and was educated in drawing and painting in Maastricht.


Ploem received his education at the University of Utrecht in the Netherlands, Harvard University and the University of Amsterdam. He has since then been employed by a number of academic institutions, including the University of Miami, Harvard, the University of Amsterdam, and the University of Leiden, where he served as a professor at the Faculty of Medicine. He also cooperated with industry, in particular in the branch of optics and concentrated on research in image analysis, participating in a project aiming to automate cancer cell recognition.

Ploem's prototype fluorescence epi-illuminators and microscopes form a part of the permanent exposition of the Dutch National Museum for the history of Science and Medicine.

Ploem described a new sub-category of digital art. Transforming digital algorithms were used to create novel pictorial handwritings for digital painting.

Ploem is Professor Emeritus at Leiden University, the Netherlands. He is a graduate of Utrecht University, the Netherlands, receiving an MD in 1972. He worked as Intern in the Broussais Hospital (Paris, France) with Professor Pasteur Valery-Radot. In 1963, Dr. Ploem was elected a Fulbright Fellow for Study at the Harvard University School of Public Health, receiving a Master of Public Health degree Cum Laude in 1954. He obtained a Ph.D. degree in 1967 from the University of Amsterdam, the Netherlands. Professor Ploem has served as visiting lecturer or professor at various universities: Dundee. Scotland; University of Florida, USA; Monash University, Melbourne, Australia; University of Beijing, China; and at the Free University of Brussels, Belgium. In 1980, he was appointed to a professorship in the Department of Cytochemistry and Cytometry at Leiden University. He retired from that position in 1992.Numerous honors and awards have been bestowed on Professor Ploem. In 1976 he was elected to the honorary Fellowship of the Royal Microscopical Society, Oxford England. In 1977 he received a Fellowship to the Papanicolaou Cancer Research Institute in Miami, Florida, and in 1979 a Fellowship to the Institute for Cell Analysis at the University of Miami, Florida. In 1982, he was the co-recipient of the C. E. Alken Foundation Award, Switzerland. In 1993, he was elected as the first Honorary Member of the International Society for Analytical Cytology. In 1993, Professor Ploem held the Erica Wachtel Medal Lecture at the British Society for Clinical Cytology meeting. In 1994, the European Society for Analytical Cellular Pathology established a Conference Keynote “Ploem” Lecture for invited scientists at its future general meetings. The International Society of Analytical Cytology invited Professor Ploem to present its inaugural “Robert Hooke” lecture. In 1995, he was invited by the Royal Microscopical Society to give the inaugural CYTO lecture.

Fluorescence microscopy[edit]

Around 1962 Ploem started work in collaboration with Schott on the development of dichroic beam splitters for reflection of blue and green light for fluorescence microscopy using epi illumination. At the time of his first publication [2] on fluorescence microscopy using epi illumination with narrow-band blue and green light, he was not aware of the development of a dichroic beam splitter for UV excitation with incident light by Brumberg and Krylova [1]. Neither was the Leitz company, Soon it became clear that excitation with narrow-band blue and green light opened optimal possibilities for the detection of the widely used immunofluorescence labels fluorescein isothiocyanate (FITC) and tetramethyl rhodamine isothiocyanate (TRITC) [3]. The use of blue and green excitation also minimized autofluorescence of tissue components an undesired effect encountered with conventional transmitted illumination with UV light. FITC could now be excited with narrow band blue light (using a band interference filter with a half width of 16 nm), close to the excitation maximum at 490 nm (long wavelength blue), with clear observation of the green fluorescence peak emission at 520 nm. Autofluorescence of tissue components was minimized (Fig. 2a, b) resulting in a high image contrast [4]. Excitation of FITC near its excitation maximum enabled such an efficient excitation that even a mercury high-pressure arc lamp, having no strong emission peak in the blue wavelength range, could be used [5]. Furthermore, epi-illumination with a green reflecting dichroic mirror enabled for the first time the excitation of Feulgen pararosaniline with the strong mercury emission line at 546 nm. In his second publication on the multi-wavelengths epi-illuminator [3], describing a Leitz prototype with four dichroic beam-splitters, Ploem could acknowledge the contribution of Brumberg and Krylova [1]. The inaccessibility of Russian research in that time period, and the absence of any major industrial development of epi-fluorescence microscopy in Russia or East Germany was the reason that Leitz and Zeiss had not been aware earlier of such a development. The possibility to introduce epi-illumination with UV light, although useful for several applications, had not been a motive for a new technological development at Leitz, since they had already excellent transmitted dark field UV excitation available. Subsequently Leitz developed a novel multi-wavelength fluorescence epi-illuminator (Leitz PLOEMOPAK) with four rotating dichroic beamsplitters for respectively UV, violet, blue and green light]. In successive generations of Leitz illuminators (containing four dichroic beamsplitters) barrier filters and a rotating turret for excitation filters were added [3,6.7]. Finally an elegant epi-illuminator was constructed by Kraft [8] containing multiple sets of a combination of an excitation filter, a dichroic beamsplitter and a barrier or emission filter, mounted together in a filter cube, also called filter block (Fig. 4). Since this illuminator permitted the filter cubes to be rapidly turned into the optical light path, multi-wavelength illumination of the same section of tissue became a practical proposition. Moreover, the four filter cubes in the illuminator could be exchanged by the user (Fig 1). Different sets of four filter cubes could be assembled, chosen from many filter cubes, containing combinations of excitation, barrier filters and dichroic beamsplitters, developed for different applications. The increasing worldwide use of routine immunofluorescence microscopy in medical diagnosis and molecular biology research could, however, profit from the new possibility @9 of epi-illumination permitting using narrow band excitation with blue and green light [9] . An important step for studies with live cells, was the design by Ploem of the first inverted fluorescence microscope with a multi wavelength epi illuminator under the microscope stage.. This provides free working stage above the microscope stage for eventual manipulation of the cells. . Following these suggestions by Ploem, Leitz also manufactured an inverted microscope with epi-illumination.

Fig. 4: Complete Leitz (Leica) filter cube (block) for fluorescence microscopy containing: excitation filter, dichroic beam splitter and emission (barrier) filter.

The Leitz (Leica) filter cube system was so efficient that now, >45 years later, similar types of filter cubes are still used by most microscope manufacturers for multi-wavelength fluorescence microscopy. This development finally led within Leica to the development of automated multi-wavelength fluorescence epi-illuminators accommodating eight filter cubes for various wavelength ranges. When switching between filter cubes, pixel shift on the computer monitor is avoided or stays below the resolution power of a 35 mm film due to a 0-pixel shift technology. This illuminator is now used for fluorescence in situ hybridisation methods (FISH) in the study of chromosomes [10]. Fluorescence microscopy of formaldehyde induced neurotransmitter CA fluorophores can be observed for the first time as blue, by using a special dichromatic mirror [11]. The independent detection of FITC (using blue excitation light) and of TRITC (using green excitation light) in the same cell uis possible with two dichromatic mirrors [12]


[1] Brumberg EM, Krylova TN: O fluoreschentnykh mikroskopopak. Zh. obshch. biol. 14: 461 (1953).

[2] Ploem JS: Die Möglichkeit der Auflichtfluoreszenzmethoden bei Untersuchungen von Zellen in Durchströmungskammern und Leightonröhren. Xth Symposium d. Gesellschaft f. Histochemie 1965. Acta Histochem. Suppl. 7: 339–343 (1967).

[3] The use of a vertical illuminator with interchangeable dichroic mirrors for fluorescence microscopy with incident light JS Ploem Z Wiss Mikrosk 68 (3), 129-142 246 1967

[4] Cormane RH, Szabo E, Hague LS: Immunofluorescence of the skin: the interpretation of the staining of blood vessels and connective tissue aided by new techniques. Br. J. Derm. 82 (Supplement 5): 26–43 (1970).

[5] JS Ploem A study of filters and light sources in immunofluorescence microscopy, Annals of the New York Academy of Sciences 177 (1), 414-429

[6] Walter F: Eine Auflicht-Fluorescenz-Einrichtung für die Routinediagnose. Leitz Mitt. Wiss. u. Techn. IV/6: 186–187 (1968).

[7] Pluta M: Handbook: Advanced Light Microscopy. Specialized Methods Vol. 2. Elsevier, Amsterdam (1989).m,

[8] Kraft W: Die Technologie des Fluoreszenzopak. Leitz Mitt. Wiss. u. Techn. IV/6: 239–242 (1969).

[9] Nairn RC, Ploem JS: Modern microscopy for immunofluorescence in clinical immunology. Leitz\ Leitz-Mitt. Wiss. u. Techn 4, 225-238 51 1969

[10] Multiple fluorescence in situ hybridization PM Nederlof, S Van der Flier, J Wiegant, AK Raap, HJ Tanke, JS Ploem, ...Cytometry 11 (1), 126-131 281 1990

[11] Ploem JS: The microscopic differentiation of the colour of formaldehyde-induced fluorescence. Prog. Brain Res. 34: 27–37 (1971).

[12] Hijmans W, Schuit HRE, Hulsing-Hesselink E: An immunofluorescence study on intracellular immunoglobulins in human bone marrow cells. Ann. N.Y. Acad. Sci. 177: 290–305 (1971).

Digital Painting[edit]

Meadow near Sauto, Pyrenees, France

Early days

Bas Ploem started painting as a small boy., While still at secondary school he attended an evening course in drawing and painting at the “Kunstnijverheidsschool Maastricht”.'

Meeting with the painters Frits and Yves Klein in Paris'

Ploem’s presence in Paris was important for his knowledge and interest in art since he could regularly visit his cousins in Paris, the painter Frits Klein and his son Yves. He visited the Kleins when Yves was making his first monochromes.

Analogue paintings

Computer image analysis for the creation of digital graphics

In the last years of his activities at the faculty of medicine at Leiden University, he concentrated on research in image analysis. He was asked to participate in a European project with the aim of automating cancer cell recognition using computer analysis. It concerned a collaborative project with the German optical company Leitz/Leica Microsystems, and the Institute for Mathematical Morphology in Fontainebleau, France. Together with a team, Professor Jean Serra at this institute had developed an image analysis method, now internationally known as ‘Mathematical Morphology’ (MM). With his experience as an analogue painter, Ploem saw the possibility of also applying the methods of mathematical morphology to the creation of digital art .

DIGITAL TRANSFORMATIONAL ART This century can be called the information century. It can be expected that fine art will profit from this technology. Since the massive advent of computers, individuals increasingly study information digitally. This has led to the study of informatics with mathematical methods and there is now an intense interaction between humans and information systems. The field of informatics encompasses many individual specializations. A group of mathematicians at the Centre of Mathematical Morphology in Fontainebleau, France, have developed a comprehensive system for image analysis using transforming algorithms. Several other programs like Adobe Photoshop 2014 and Corel PaintShop Pro X5/X7 permit the use of transforming algorithms . Corel PaintShop Pro enables effective algorithms for erode and dilate. Leonardo da Vinci wrote already a paper on image transformation.

Is seems unavoidable that in the present century with informatics influencing our daily lives, the digital technologies associated with informatics will also be applied by some artists to the analysis and the creation of art.

Transformation of forms, changing one form into another, has always been performed by artists. When Vincent van Gogh in his later works observed round or oval leaves, he changed (transformed) these leave shapes in his brain and painted images of trees with a type of curved brackets. Many other painters as well have in their minds changed the objects they see in some other shape, before painting them on canvas or paper. In this way they created their own characteristic pictorial or handwriting.

At the International Symposium on Mathematical Morphology in Amsterdam (1998), J.S Ploem presented a paper on the creation of computer graphics with Mathematical Morphology, using for the first time, the transforming algorithms from the Fontainebleau group for the creation of digital art. (Mathematical Morphology and its Applications to Image and Signal Processing, Heijmans and Roerdink (Eds), Kluwer. 1098: page 355:) Ploem described and developed methodology for the creation of transformational digital art. The creative tool in classical digital art is often the use of the computer mouse. With digital transformational art, however, mathematical algorithms are the main creative tools and not the drawing by a computer mouse. The term transformational art has already been used earlier for some interesting categories of art. But in this website this term is again proposed for the special discipline of digital image transformation, following the earlier use of this term for image transformation, as described by Leonardo da Vinci in 1505. Transformational art, as described in this chapter, requires always a source image or drawing that has to be changed (transformed) into a target image. Algorithmic, mathematical and fractal art do not need source images, since they can create an entirely new image on their own. Ploem used for image transformation the QWIN program from LEICA, based on the French Mathematical Morphology program. The QWIN program can perform transformations on binary images and on gray scale images. A binary image is obtained by thresholding the gray scale image. A range of intensity (gray) levels in the image can be chosen for detection. It can be transformed with the various “amend” instructions from QWIN such as : erode, dilate skeleton, ultimate skeleton, opening, closing, prune . “skiz“ etc. The watershed transformation is performed on grey scale images. It is a rather aggressive algorithm that can damage some morphology in a picture. It is very effective in creating totally new pictorial handwritings. For technical details one could consult a website on transformational art ( The pictorial handwriting is an essential component of a painting. Using digital transformational algorithms, novel pictorial handwritings can be created. Such handwritings are very difficult or almost impossible to make with hand methods. By means of modern computer technology and mathematical morphology programming new pictorial handwritings can be generated. It is, however, often impossible for a digital artist to predict the pictorial results in programming with transforming algorithms. Due to the randomization effect of some algorithms, very variable patterns will be created. The handwritings produced do not always show the right color palette for an intended painting. If the digital artist would use the Photoshop color palette construction program, he would obtain a quite large variation of colored results. Only a few results may be of artistic interest. Full interaction by a digital artist is therefore needed to select only the useful results. Choosing from all these images the right image is a time consuming procedure and requires considerable (artistic) talent.

In the valley of the Eyne, Pyrenees

Mountain flowers as the first topic for digital image analysis

His first digital graphics of nature scenes were shown in his exposition at a regional art centre in the Pyrenees (Ossega, June, 1997).

Scientific interest in computer graphics created with mathematical morphology

As he was probably the first person to systematically use mathematical morphology for the creation of digital art, Ploem’s work attracted international attention and he was invited as a plenary speaker at an international mathematical conference in Amsterdam in 1998 to explain his new type of digital art. He also received an invitation to show his art work in an exposition at this conference. The organisers of this meeting asked Ploem to write a chapter on his novel technique for digital art in a book (Kluwer, ISBN 0-7923-5133-9) that was published on the occasion of this meeting.

Exposition at universities

He was invited for a symposium on ‘Art et Science’ at the University of Caen, France (April, 2001). At the art exposition connected with this symposium, he presented 6 digital graphics that were dominated by chaotic transformations of rock art themes. A similar invitation was made by the University of Basel in Switzerland (April, 2002).

David and Goliath (multiple image transformations of a romanesque painting in the Sant Climent church in Taull)

Portraits and architecture

External links[edit]