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Home / Palaeobotanical Tools / Focus stacking (Photography, Extended Depth of Field)

Preparation and Conservation
Managing Fossil Collections
Transfer Technique
Palynological Preparation Techniques
Cellulose Peel Technique
Latex Casts
Photography and Scanning
Fluorescence Microscopy and Fluorescence Microspectroscopy
Scanning- (SEM) and Environmental Scanning Electron Microscopy (ESEM)
Digital Cameras on the Microscope
Cameras With Focus Bracketing or Built-In Focus Stacking
Superresolution (SR)
High Dynamic Range Imaging (HDR)
Image Processing
Imaging Fossils Using UV-Light (Black-Light Photography)
Transmission Electron Microscopy (TEM)
Microtomography (CT Scanning, XTM) including Synchrotron X-ray Tomographic Microscopy (SRXTM)
Raman Spectroscopy
Writing, Translating and Drawing

! Panorama Creation and Stitching Tools@

Focus stacking (Photography, Extended Depth of Field)

! A. Agarwala et al.: Interactive Digital Photomontage. In PDF. See also here.

! Peter Bargh, YouTube: Focus Stacking - a macro photography technique. The use of free Combine ZM software or Helicon Focus software.

R.S. Barclay et al. (2013): New methods reveal oldest known fossil epiphyllous moss: Bryiidites utahensis gen. et sp. nov.(Bryidae). In PDF, American Journal of Botany, 100: 2450-2457.
Using the zstacking software package Automontage (Syncroscopy, Cambridge, UK).

R. Berdan (2017), The Canadian Nature Photographer:
! Focus Stacking comparing Photoshop, Helicon Focus and Zerene.
"... For stacking images once in a while, I think that Photoshop does a satisfactory job but lacks the ability to tweak the stacking process. Helicon was the easiest software to use, supports RAW files, offers features to optimize stacking and creates 3D files from the stacks. Zerene also created nice stacks and stereo images, though its lack of RAW file support and high price makes the other programs more attractive ...".

! A. Bercovici et al. (2009): Improving depth of field resolution for palynological photomicrography. PDF file, Palaeontologia Electronica, Vol. 12.

V. Blagoderov et al. (2012): No specimen left behind: industrial scale digitization of natural history collections. In PDF, Zookeys, 2012, (209): 133-146.

Bradley, A.P., Wildermoth, M. and Mills, P. (2005): Virtual Microscopy with Extended Depth of Field. Abstract, Digital Image Computing: Techniques and Applications (DICTA'05), Cairns, Australia.

! J. Brecko et al. (2014): Focus stacking: Comparing commercial top-end set-ups with a semi-automatic low budget approach. A possible solution for mass digitization of type specimens. In PDF, ZooKeys, 464: 1-23. With a comparison of different focus stacking software packages (PDF page 19). Excellent!
See also here.
"Based on the results presented we can conclude that the Canon-Cognisys set-up as we currently use it delivers results that are equal, or even better, than high-end approaches. This merely is due to the simple lighting set-up, the high resolution, and the low noise delivered by the Canon DSLR".

Matt Brett, Digital Photography School, Canada: An Introduction to Focus Stacking.

M. Bridge (2016), SLR Lounge: Is Helicon Focus The Best Focus Stacking Software? {Review}

Cambridge in Colour (by Sean McHugh). This is an online learning environment where photographers can come to improve their skills. Focus Stacking & Depth of Field.

Capture the Atlas:
Focus Stacking in Photography – Step by Step Tutorial.

Canon Digital Photography Forums: Which Focus Stacking Software?

Andrew Childress, Envatotuts: Focus Stacking Made Easy with Photoshop.

Creative Cloud:
Focus Stacking in Photoshop. In Geman. Focus stacking. Search results (in German).

Dennis Goulet Photography: Focus Stacking - Part 3, What Works, What Doesn´t. (in German):
Search results: Focus stacking

Ed Dozier (2016): Focus Stacking With Combine ZM.

Dreessen Informatik:
Focusstacking mit CombineZP.
Makro stacking mit CombineZP Anleitung. In German.

Explora (B & H Foto & Electronics Corp., New York):
Photo-Editing Tutorial: Focus Stacking for Macro Photography (by Shawn C. Steiner).

! Extreme-Macro (by Johan J Ingles-Le Nobel).
A free online guide to macro photography, including focus stacking, macro techniques, lighting, software and more.
Worth checking out: The focus stacking workflow.

H. Failmezger et al. (2013): Semi-automated 3D Leaf Reconstruction and Analysis of Trichome Patterning from Light Microscopic Images. In PDF, see also here.

U. Fischer (2016):
Focus Stacking leicht gemacht. In German.

flickr: Macro Viewers / Discussion: Focus stacking Tutorial. Using the freeware programme combinez5 and links to other tutorials. See also here.

FocOz: FocOz Software. This program is comparing different frames, selecting image elements of high definition from them all and rejects whatever is blurred. (in German). Focus-Stacking. In German.

Daniel Glasner et al.: Super-Resolution From a Single Image.

Stefan Gross, Traumflieger: Photoshop-Workshops, Mehr Schärfentiefe mit Photoshop. The Photoshop procedure, a tutorial (in German).

! Alan Hadley, U.K.: CombineZ Movie CZM).
This expired link is available through the Internet Archive´s Wayback Machine. For download see also here.
The purpose of this free program is to increase depth of focus by combining pictures, but each picture should be in focus at a different depth into the scene. The program is especially useful to microscope users (e.g. palynomorphs) and macro photographers. Excellent! CZM is the latest version, it can process movies, take pictures, and has more macro commands but still has all of the functionality of the former CombineZ5. Still available:
CombineZ5. It works with older versions of windows, but is not maintained anymore. See also:
Yahoo Group combinez,
Forum discussions (Digital Grin), or
Wikipedia, CombineZM.
Tutorials are here (flickr), and there (Brian Valentine).

J.T. Haug et al. (2011): Autofluorescence imaging, an excellent tool for comparative morphology. In PDF, Journal of Microscopy.

Helicon Soft Ltd., Kharkov, Ukraine: Helicon Focus. This is a shareware program that creates a completely focused image from several partially focused images by combining the focused areas. You can download and use the program for evaluation purposes for 30 days. Helicon Focus is designed for macrophotography, microphotography (fluid inclusions!) and hyperfocal landscape photography to cope with the shallow depth-of-field problem.
See also Video Tutorials.

Helicon Soft: Helicon Remote. Helicon Remote software allows you to tether a Canon or Nikon DSLR camera to a computer or a mobile device and automates focus (DoF) bracketing, exposure bracketing (HDR), and time lapse photography.

! John Hollenberg, Digital Outback Photo: Workflow Technique 072. Review of Helicon Focus and CombineZ5 software. Excellent!

! Neal Immega, Houston Gem and Mineral Society: how2doit Articles. Go to:
Extended Focus in Macro Photography (PDF file). A CombineZ5 tutorial. See also here.

Infognition: What is super-resolution?

! J.J. Ingles-Le Nobel, Extreme Macro: Focus Stacking.
And here an older version: Macro Stacking.

O. Jaurich (?): Mikroskopie und mikroskopische Bildverarbeitung, Nachbearbeitung der aufgenommenen Bilder. About CombineZ5 (in German).

Sarah Johnson,
Digital Photography Basics.
This digital photography guide will cover every aspect of digital photography to make digital photography easier, and help you improve your photography skills.

! G. Kontogianni et al. (2017): Enhancing close-up image based 3D digitisation with focus stacking. Free access, isprs (The International Society for Photogrammetry and Remote Sensing).

Biomedical Imaging Group, EPFL, Lausanne, Switzerland: Extended Depth of Field. Extended depth of field is an image-processing method e.g. to obtain in focus microscopic images of 3D objects. Free and easy-to-use focus stacking software.

Rik Littlefield, Richland, WA: An Introduction to Extended Depth of Field Digital Photography (from News of the Lepidopterists' Society, Summer 2005) and
Extended Depth of Field Photography of Insects. The software used here was CombineZ5. See also: Extended Depth-Of-Field Using Panorama Tools.

Jay Longson et al. (2010): Adapting Traditional Macro and Micro Photography for Scientific Gigapixel Imaging. PDF file, Proceedings of the Fine International Conference on Gigapixel Imaging for Science, November 11–13.
See also here.

Max Lyons, TAWBAWARE. TawbaWare is home to software, photographs, and writings. Go to:
TuFuse Pro. Graphical Exposure and Focus Blending Software.

Makrotreff: Focus Stacking leicht gemacht!. In German.

Media Cybernetics Inc.: Image-Pro Plus. The Extended Depth of Field function in Image-Pro Plus provides you with the ability to display a stack of images as a composite, building in effect a three dimensional image.

! J.E.J. Mertens et al. (2017): The use of low cost compact cameras with focus stacking functionality in entomological digitization projects. Open access, Zookeys, 712: 141–154.
Images of a professional setup were compared with the Olympus Stylus TG-4 Tough, a low-cost compact camera with internal focus stacking functions.

G. Montesanto (2015): A fast GNU method to draw accurate scientific illustrations for taxonomy. Zookeys, 515: 191–206.

National Institutes of Health, Department of Health and Human Services, Bethesda, Maryland (program developed by an employee of the Federal Government in the course of his official duties): ImageJ. ImageJ is a public domain Java image processing program. It runs, either as an online applet or as a downloadable application, on any computer with a Java 1.4 or later virtual machine. It can display, edit, analyze, process, save and print 8-bit, 16-bit and 32-bit images (including TIFF, GIF, JPEG, BMP, DICOM, FITS and "raw"). ImageJ supports "stacks" and standard image processing functions such as contrast manipulation, sharpening, smoothing, edge detection and median filtering. It can calculate area and pixel value statistics of user-defined selections, measure distances and angles and it can create density histograms and line profile plots.

Nikon Professional Services (NPS). This is a Nikon organization designed solely to assist qualified, full-time professional photographers:
Focus Shift Photography.
! Technical Solutions, D850 TIPS: Focus Stacking (Focus Shift Photography). In-Camera Focus Stacking.

! Optronics: Rincon. An image measurement and processing software. Rincon can combine a stack of images captured at different levels of focus and combine them into a single in-focus image. Macro photo stacking software comparison.

PhotoAcute, Austin, Texas, USA: Superresolution FAQ.

! How To Focus Stack Images In Photoshop

! PICOLAY (from PICture OverLAY, by Heribert Cypionka, Institut für Chemie und Biologie des Meeres, Universität Oldenburg, Germany). This is a freeware program for Windows computers designed for the extraction of improved images from digital picture series. Excellent!

! R. Pintus et al. (2019): State-of-the-art in Multi-Light Image Collections for Surface Visualization and Analysis In PDF, Computer Graphics Forum, 38. See also here.
"... Multi-Light Image Collections (MLICs), i.e., stacks of photos of a scene acquired with a fixed viewpoint and a varying surface illumination
[...] We [...] review techniques that improve object understanding by using illustrative approaches
[...] We also review how these methods are applied in several, main application domains, and what are the available tools to perform MLIC visualization and analysis. We finally point out relevant research issues, analyze research trends, and offer guidelines for practical applications ..."

! J. Piper (2010), go to PDF page 190: Software-Based Stacking Techniques to Enhance Depth of Field and Dynamic Range in Digital Photomicrography (PDF file). In: T.D. Hewitson, I.A. Darby (eds.), Histology Protocols, Methods in Molecular Biology 611.

! Pro Photo: Focus Bracketing und Focus Stacking: Automatische Fokus-Aufnahmereihen. About some new camera types producing photographs with extended depth of field. In German.

E. Reeves et al. (2023): Historic palaeobotanical collection reveals in situ microspores and pollen from Early Carboniferous (Tournaisian) ovules from the Ballagan Formation of Scotland. Free access, Review of Palaeobotany and Palynology, 308.
"... The resultant 67 photomicrographs were photo-stitched into one large image
[...] Single microspores and pollen within the ovule were photographed under oil at ×100 using multiple images at different focus depths and then Z-stacked ..."

Reindeer Graphics, Asheville, NC: Optipix software.

J.B. Riding and M.J. Head (2017): Preparing photographic plates of palynomorphs in the digital age. Palynology, 42. See also here (in PDF). Worth to read:
Chapter 5.5.2 (on PDF page 6): "Multiple images versus image stacking".

Ruedi Rottermann et al. (2010): How Sharp Images Are Formed - Depth of Field in Microscopy. About the Multifocus module of the Leica Application Suite (LAS).

Saphicon: AllFocus. The AllFocus software aligns images in the Z-stack by shifting and scaling them and creates a composite image by choosing pixels from the best-focused plane.

! M. Schädel, University of Tuebingen: Multi Light Imaging of small compression fossils – proper tools and a neat shortcut. In PDF, Conference poster: The Palaeontological Association 67th Annual Meeting, University of Cambridge, UK.
About Multi Light Imaging with focus stacking or panoramic stitching.

H.A. Schmid, Pilze Forum: Coolpix-Lupenmakro/CombineZ5 (Anleitung). About CombineZ5 (in German).

! Stefan Schorn, Mineralienatlas, Germany: Digitale Mehrebenen-Fotografie (DMF) / Digitale Fokuserweiterung. A tutorial, including a list of further focus stacking software programs (in German). See also:
! CombineZ5 / CombineZM. A short tutorial.
This expired link is now available through the Internet Archive´s Wayback Machine.

SCM Altivid, Berlin: Mikroskopkamera (in German). A low-cost camera with extended depth of field.

! P.A. Selden (2014): A workflow for digital photography of fossil specimens. In PDF, The Geological Curator, 10: 93-97.

Traumflieger Fotographics (Stefan Gross, Hamburg): Camera tests and descriptions, reports and a forum, especially about marophotography (in German).
! So funktioniert Fokus-Stacking!
Extrem-Makros jenseits von 1:1! (Video report).

Brian Valentine: Wonderful Photos: How to Increase your Depth of Field by Focus Stacking. A tutorial on using the freeware CombineZM (by Alan Hadley, U.K.).
This expired link is still available through the Internet Archive´s Wayback Machine.

Wikipedia, the free encyclopedia:
! Focus stacking.
Category:Photographic techniques.
Category:Free photo software.

Kurt Wirz, Switzerland:
Focus Stacking. In PDF. A gallery of beautiful focus stacking images. See especially:
! Making of.

Bruce Wunderlich, Digital Photography School:
A Beginner’s Guide to Focus Stacking.

D. Zheng et al. (2018): A Late Cretaceous amber biota from central Myanmar. In PDF, Nature Communications.
"To get the clear 3D structures, all images were taken by using digitally stacked photomicrographic composites of ~40 individual focal planes".

Zerene Stacker (by Zerene Systems, LLC., Rik Littlefield). This is is a "focus stacking" program designed specifically for challenging macro subjects and discerning photographers. The new programme Zerene Stacker is available for a free trial period.

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