Everspark

The super-resolution microscopy buffer that offers multicolor, long-term and stable fluorescence imaging:
prepare your samples one day, and image them every day for weeks

 Features      Protocol    Story
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Odoo • Image et Texte


What is it for? Use it if you want to decouple the preparation of your samples and their imaging.  For instance if you need to prepare your samples in your lab, and image them in your core facility 3 to 4 weeks later.
Use it if you want to image with red, far red and green dyes (eg. AF488). Use different colors either at the same time or one after the other.

Designed by Karine Monier, Arnaud Favier and Christophe Place 

Reviewed by Pierre Bon, Ingrid Chamma, Benjamin Compans, Laurence Dubreil, Beatrice Durel, Tamas Gajdos, Sébastien Mailfert, Céline Malleval, Paolo Marchi, Delphine Muriaux, Jonny Nixon-Abell

Powered by the Eternity technology published in Scientific Reports 


The protocol of use with a glass-bottom dish

 
 
The protocol of use with a depression slide

 
 


Features

High stability over time for repeated measurements

One mounted sample is stable over 3 to 4 weeks and can be imaged multiple times

Multicolor (incl. green)

 
Compatible with green, red & far red dyes (AF488, AF647, CF647, DL550, CF568, DL650, CF680, SulfoCy5) and mEos2

Ready to use


Can be used directly from the vial

Up to 6 months performance

Individualized packaging for optimal longevity

Techniques


PALM
dSTORM
HILO, TIRF pattern

Option: customizable


Choose the MEA concentration, pH, optical index, or even additives

  Community ongoing tests

The community is currently testing those conditions:
Multiconditions at the same time or sequentially: High Content Screening / High Troughput

The community disclosed that Everspark does not work 100% efficiently with organs and tissues  because there is most of the times a bit of remaining oxygen.

And we would love to know if somebody is testing PALM + STORM at the same time, as well as multiple conditions at once using Stencell !

  Gallery

The recipe to prepare a good sealing

 
 

  Simplified protocol

Storage conditions: 

  • Everspark has to be stored in the fridge at 4°C upon receiving the product. Do not freeze it.

  • Open the tube just before mounting to limit oxygenation

  • Store the slides at 4°C and in the dark before repeating dSTORM acquisition on the same slide.

Use method:

  • Rinse the coverslip with sterile water

  • Discard water in excess

  • Prepare sealant: 1 volume of sealant catalyst + 1 volume of sealant basis in a micro-dish – do not mix

  • Open the Everspark tube and pipette right away the desired volume onto a Wilco glass bottom dish

  • Flip the coverslips face towards the slide on the cavity filled with Everspark without making bubbles

  • Absorb excess of Everspark buffer on the sides with a kimwipe

  • Mix sealant

  • Seal quickly with the reconstituted sealant

  • Wait for the sealant to polymerize on the slide before moving the slide

  • Discard the open tube of Everspark buffer (cannot be used for another slide, once already opened)

  • Gently clean the coverslip with 70% ethanol

  • Perform regular dSTORM acquisition (best to wait a couple of hours)

  • Repeat dSTORM acquisition 24 hours up to 3 weeks on the same slide providing no air bubbles were present

  • Store the slides at 4°C and in the dark before repeating dSTORM acquisition on the same slide (up to 3 weeks)

Note 1: Blinking events have been validated with Alexa-fluor 647, DL650, DL550 and CF568 (more to come)

Note 2: Blinking is detected for several weeks without loss of events or photons per event

Note 3: Mounting can also be performed in various microdishes by adjusting the Everspark volume (Each vial contains 450 µl)

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Everspark and us

"We designed the Everspark buffer to facilitate sample preparation for dSTORM super-resolution imaging. Indeed, an oxygen-free buffer containing a thiol reducer is central to promote fluorophore blinking and thus single molecule detection, required for dSTORM imaging. With classical buffers, such as the Glox buffer, fluorophore blinking lifetime is limited to a few hours. With Everspark buffer, it is now possible to increase sample lifetime to several weeks. We demonstrated that a calibrated sample mounted once with the Everspark buffer can be submitted to several rounds of dSTORM imaging to acquire and reconstruct a structure reproducible in coverage and precision, over a period of two months."

Contact the team about their technology.

Odoo • Text and ImageKarine Monier - Christophe Place -  Arnaud Favier
Odoo • Image and Text Esther Graudens
New projects at Idylle

"When I was working on long manipulations, I dreamed of being able to stop at certain crucial stages. Everspark is the ideal answer to the problem of many researchers who prepare their samples in a hurry simply because they need to image them right away. The Everspark buffer eliminates this tension: they take the time to image whenever they want, with complete peace of mind. They can also make acquisitions over several days. This will really change the way people prepare their experimentations, and that's exactly what I like about my job!
In terms of production, I am very proud of the way we have managed to set up a just-in-time production process in our labs in record time. And when researchers ask us to introduce variants, our production teams are able to build and produce very quickly their customized buffer to test it".

Contact us about your sample preparation workflow

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