Joachim Goedhart

@joachimgoedhart

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Joachim Goedhart@joachimgoedhart· 28d

It certainly appears to be so (in C.elegans) source: academic.oup.com/genetics/art...

fluorescence images of GRP1 in c.elegans

Joachim Goedhart@joachimgoedhart· 28d

We also see nuclear accumulation for Grp1 - it's probably caused by the polybasic stretch near the end of the domain, acting as nuclear localization signal.

SPFKIPEDDGNDLTHTFFNPDREGWLLKLGGRVKTWKRRWFILTDNCLYYFEYTTDKEPRGIIPLENLSIREVEDPRKPNCFELYNPSHKGQVIKACKTEADGRVVEGNHVVYRISAPSPEEKEEWMKSIKASISRDPFYDMLATRKRRIANKK*

Joachim Goedhart@joachimgoedhart· 28d

Cheek epithelial cells from (this) human stained with Acridine Orange #FluorescenceFriday

Fluorescence image of cheek epithelial cells stained with Acridine Orange

Joachim Goedhart@joachimgoedhart· 29d

The introduction of "Inventing temperature" by Hasok Chang starts well (actual text in the alt-text). #InventingTemperature

However, the cutting edge is not all there is to science, nor is it necessarily the most valuable pan of science. Questions that have been answered are still worth asking again, so you can understand for yourself how to arrive at the standard answers, and possibly discover new answers or recover forgotten answers that are valuable.

Joachim Goedhart@joachimgoedhart· Jul 15

In preparation for a practical course I measured the nuclear area of several human cell types, and the differences are significant!

Graph showing the nuclear area of human cheek, endothelial and HeLa cells

Joachim Goedhart@joachimgoedhart· Jul 1

Version 1 has low chemical stability, may that explain why?

Joachim Goedhart@joachimgoedhart· Jul 1

A High-Affinity Nanobody Recognizing mNeonGreen Enables Versatile Biochemical, Cellular, and in vivo Applications. by Steffen Frey and team: www.biorxiv.org/content/10.6...

$mNeonGreen Selector: a pulldown resin for mNeonGreen-fusion proteins. (A) Sketch of mNG Selector binding mNG fusion proteins. (B, C) Immunoprecipitation from E.coli lysate in the presence (B) or absence (C) of target protein (mNG-Halo fusion, arrowhead). Proteins were eluted in boiling SDS sample buffer. mNG Selector enables highly specific one-step capture of mNG-tagged proteins. (D) Binding assay to evaluate the cross-reactivity of the mNG Selector to other recombinant fluorescent proteins. Shown is a figure of the equilibrium bound fraction of each indicated fluorescent protein prior to buffer washing. (E) Compatibility of mNG Selector to various reagents applied during binding (top panel) or during washing steps (lower panel). mNG fluorescence is lost in the presence of 2 M Gua-HCl. The symbols below each row indicate binding efficiency: ++: strong binding, + binding, -: no binding$

Joachim Goedhart@joachimgoedhart· Jun 23

Navigating online systems to achieve a simple thing (sending a package) often feels like playing King's Quest

Screenshot of King's Quest with king Edward on a throne

Joachim Goedhart@joachimgoedhart· Jun 20

Soccer field unable to choose between dark mode and light mode.

Screenshot of soccer match, where half the field is in the shade

Joachim Goedhart@joachimgoedhart· Jun 12

Fluorescence Lifetime Imaging in Plants: Practical guidelines for multiplexing, label-free imaging and data analysis by Thomas Ott and team: www.biorxiv.org/content/10.6...

Figure 2. Live-cell multicolour lifetime imaging with two pairs of fluorescent reporters in different spectral windows. A) Two green- (EGFP and NowGFP) and two red- (mCherry and mScarlet) emitting FPs with large lifetime differences (Δτ) were targeted to four different compartments of N. benthamiana leaf epidermal cells (schematic). PM = plasma membrane; Chl = chloroplasts; N = nucleus; ER = endoplasmic reticulum. B) Intensity-only and FastFLIM confocal images of an epidermal cell showing the discrimination of two fluorescent reporters within the green and red channel based on lifetime values (rainbow bar). C) Identification of the fluorescent reporters and mapping of their subcellular distribution based on the Phasor fingerprint. The white dashed box indicates the region shown in the close up in (D)

Joachim Goedhart@joachimgoedhart· Jun 10

Programming cell behavior with synthetic protease-activated receptors by Alice Y Ting and team: www.biorxiv.org/content/10.6...

Figure 1 SynPARs integrate pericellular protease activity to drive defined outputs. (A) Synthetic protease activated receptors
(SynPARs) are comprised of a protease-cleavable autoinhibitory domain tethered to a GPCR. Cleavage of the input module relieves
inhibition, allowing either an exogenous agonist (Chemo-SynPAR, top) or a masked peptide agonist (Auto-SynPAR, middle) to activate
the GPCR. This conditional activity can be coupled to a variety of outputs (bottom) including light/BRET-gated transgene expression
(SynPARTF), endogenous G-protein coupling (SynPARG), and real-time fluorescence (SynPARFL).Luc, luciferase. TEVp, TEV protease.
TF, transcription factor. cpFP, circularly permuted fluorescent protein.

Joachim Goedhart@joachimgoedhart· Jun 8

Find the latest version of SuperPlotsOfData here: uva-nl.shinyapps.io/SuperPlotsOf... It supports multi-file upload and now has the option to jitter the large dots (mean of median of replicates) No jitter Jitter

Superplot-style plot without jitter of the means of replicates - not all means visible due to overlap

Superplot-style plot with jitter of the means of replicates - all means visible!

Joachim Goedhart@joachimgoedhart· Jun 7

Topology-dependent FRET efficiency in living cells via N-C swapping of fluorescent protein fusions by Takayuki Nakajima and team: www.biorxiv.org/content/10.6...

Figure 3. Sensitized emission FRET measurements in living cells

Joachim Goedhart@joachimgoedhart· May 20

A Spectrum of Possibilities: A Systematic Evaluation of Fluorescent Proteins in Cyanobacteria by Anna Behle and team: www.biorxiv.org/content/10.6...

Optimal signal-to-background ratios plate reader of each FP strain

Joachim Goedhart@joachimgoedhart· May 19

Combining biosensor imaging data with an animated plot of the quantified intensity, based on a blog I wrote earlier: thenode.biologists.com/crafting-plo... In this version I used a dark theme

Joachim Goedhart@joachimgoedhart· May 19

Allosteric Biosensors Unravel GTPase-Effector Feedback by Klaus Hahn and team: www.biorxiv.org/content/10.6...

Schematic of AlloRac1 design: circularly permuted GFP (cpGFP) is inserted into the allosteric loop such that its fluorescence is coupled to Rac1 activity

Joachim Goedhart@joachimgoedhart· May 18

RhoG, Rac1 and Cdc42 cooperation in cell protrusion revealed by multiplexed optogenetics and biosensor imaging by Klaus M. Hahn and team: www.biorxiv.org/content/10.6...

Joachim Goedhart@joachimgoedhart· May 18

Electroporation-mediated delivery of protein biosensors for metabolic imaging in differentiated myotubes by Satoshi Arai and team: www.biorxiv.org/content/10.6...

Calcium and ATP imaging of differentiated myotubes with Tq-Ca-FLITS, a fluorescence lifetime imaging microscopy (FLIM)–based Ca2+ biosensor and MaLionG, an intensity-based ATP biosensor.

Joachim Goedhart@joachimgoedhart· May 18

Green genetically encoded IP3 biosensor for hierarchical analysis of its signaling network by Tetsuya Kitaguchi and team: www.biorxiv.org/content/10.6...

Figure 1. Schematic drawing of Green iPenguins.

Joachim Goedhart@joachimgoedhart· May 13

A CandiChrome toolkit for multicolor labeling of Candida cells by Richard J Bennett and team: www.biorxiv.org/content/10.6...

Joachim Goedhart@joachimgoedhart· May 6

Mirror mirror on the wall, which image is most natural? journals.biologists.com/jcs/article/...

Image transformations caused by standard microscope elements.

Joachim Goedhart@joachimgoedhart· May 6

Improved sensors for fructose-1,6-bisphosphate enable in vivo imaging of glycolysis by Alison Tebo and team: www.biorxiv.org/content/10.6...

Figure that summarizes the effort to produce HyLight2. It shows a structural model, results from the screens, the screening setup and spectra.

Joachim Goedhart@joachimgoedhart· Apr 24

📣 New blog-post: box plots for low n is a bad idea. Here's why and a nice fix in R/ggplot2 when you have mixed numbers of observations: thenode.biologists.com/unboxing-dat...

A plot with four conditions, and only two box plots

Joachim Goedhart@joachimgoedhart· Apr 23

LOL, was that on purpose @andreacaldarola.bsky.social ?

Text reading: I did a spelling check and mixed some typos (no assurances that I got everything!)

Joachim Goedhart@joachimgoedhart· Apr 21

TIL "dynamic range" is used for two orthogonal biosensor properties. Which one is correct?

Two plots, showing a biosensor response. Both report a 'dynamic range', one referring to the biosensor affinity and one referring to the optical signal.

Joachim Goedhart@joachimgoedhart· Apr 20

There's an update of #EZreverse, it is much quicker and supports drag-and-drop joachimgoedhart.github.io/EZreverse.html

Joachim Goedhart@joachimgoedhart· Apr 17

The original work says:

Joachim Goedhart@joachimgoedhart· Apr 17

For #FluorescenceFriday I'm sharing some promotional material (for education) made by Anna Pietraszewska

SCIENCE written with fluorescent bacteria

Joachim Goedhart@joachimgoedhart· Apr 13

All the data presented in the preprint, and the code to process the data is available (links included in the preprint). The data analysis should be completely reproducible, allowing to go from calibration data to Ca2+ concentration traces.

Curve showing the dynamics of intracellular Ca2+ concentrations in HeLa cells

Joachim Goedhart@joachimgoedhart· Apr 13

All FLIM measurements were done on the @leicamicrosystems.bsky.social Stellaris8, so it is partially a manual on how to operate that specific machine.