Autofluorescence generation and elimination: a lesson from glutaraldehyde

Lee, Kwahun; Choi, Sungmoon; Yang, Chun; Wu, Hai‐Chen; Yu, Junhua

doi:10.1039/c3cc40799c

Cited by 71 publications

(73 citation statements)

References 23 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…21 It has been shown that auto-fluorescence occurs from the chemical treatment of APTES and glutaraldehyde. 22 In our case, we were able to confirm the phenomenon was specific to glutaraldehyde and APTES ( Fig. SI 1a), and use it to show that it only occurs on PDMS in the presence of these chemicals ( Fig.…”

Section: Resultssupporting

confidence: 58%

Integrated Micropillar Polydimethylsiloxane Accurate CRISPR Detection (IMPACT) System for Rapid Viral DNA Sensing

Bao

et al. 2020

Preprint

View full text Add to dashboard Cite

A fully Integrated Micropillar Polydimethylsiloxane Accurate CRISPR Detection (IMPACT) system is developed for viral DNA detection. This powerful system is patterned with high-aspect ratio micropillars to enhance reporter probe binding. After surface modification and probe immobilization, CRISPR Cas12a/crRNA complex is injected into the fully enclosed system. With the presence of double-stranded DNA target, the CRISPR enzyme is activated and non-specifically cleaves the ssDNA reporters initially immobilized on the micropillars. This collateral cleavage releases fluorescence dyes into the assay, and the intensity is linearly proportional to the target DNA concentration ranging from 0.1 to 10 nM. Importantly, this system does not rely on traditional dye-quencher labeled probe thus eliminating the fluorescence background presented in the assay. Furthermore, our one-step detection protocol is performed at isothermal conditions (37°C) without using complicated and time-consuming off-chip probe hybridization and denaturation. This miniaturized and fully packed IMPACT chip demonstrates rapid, sensitive, and simple nucleic acid detection and is an ideal candidate for the next generation molecular diagnostic platform for point-of-care (POC) applications, responding to emerging and deadly pathogen outbreaks.

show abstract

Section: Resultssupporting

confidence: 58%

Integrated Micropillar Polydimethylsiloxane Accurate CRISPR Detection (IMPACT) System for Rapid Viral DNA Sensing

Bao

et al. 2020

Preprint

View full text Add to dashboard Cite

show abstract

“…It is well known that glutaraldehyde, the most used fixative in EM, should be avoided when fluorescence imaging needs to be performed. Glutaraldehyde-fixed samples, indeed, exhibit high levels of autofluorescence when exposed to near ultraviolet light, which decreases the image contrast when other fluorescence signal is detected [2,6,7]. Conversely, formaldehyde is a much less efficient fixative and is not able, when used alone, to guarantee a satisfactory ultrastructure conservation for EM analysis [2,4].…”

Section: Introductionmentioning

confidence: 99%

Effects of fixatives on myelin molecular order probed with RP-CARS microscopy

et al. 2020

View full text Add to dashboard Cite

When live imaging is not feasible, sample fixation allows preserving the ultrastructure of biological samples for subsequent microscopy analysis. This process could be performed with various methods, each one affecting differently the biological structure of the sample. While these alterations were well-characterized using traditional microscopy, little information is available about the effects of the fixatives on the spatial molecular orientation of the biological tissue. We tackled this issue by employing Rotating-Polarization Coherent Anti-Stokes Raman Scattering (RP-CARS) microscopy to study the effects of different fixatives on the myelin sub-micrometric molecular order and micrometric morphology. RP-CARS is a novel technique derived from CARS microscopy that allows probing spatial orientation of molecular bonds while maintaining the intrinsic chemical selectivity of CARS microscopy. By characterizing the effects of the fixation procedures, the present work represents a useful guide for the choice of the best fixation technique(s), in particular for polarisation-resolved CARS microscopy. Finally, we show that the combination of paraformaldehyde and glutaraldehyde can be effectively employed as a fixative for RP-CARS microscopy, as long as the effects on the molecular spatial distribution, here characterized, are taken into account.

show abstract

“…17). 8 Complete, high-fidelity expansion in tissue required a lower MA-NHS concentration than in cultured cells (1 mM for 60 min), presumably due to physical differences in the specimens. We therefore advise validation (through correlative imaging pre- and post-expansion) and possible optimization of these procedures or their variations before applying them to uncharacterized specimens which may have different properties.…”

mentioning

confidence: 99%

Expansion microscopy with conventional antibodies and fluorescent proteins

et al. 2016

View full text Add to dashboard Cite

Expansion microscopy is a recently introduced technique in which fluorophores on fixed specimens are linked to a swellable polymer that is physically expanded to enable super-resolution microscopy with ordinary microscopes. We have developed and characterized new methods for linking fluorophores to the polymer that now enable expansion microscopy with conventional fluorescently-labeled antibodies and fluorescent proteins. Our methods simplify the procedure, expand the palette of compatible labels, and will aid in rapid dissemination of the technique.

show abstract

Autofluorescence generation and elimination: a lesson from glutaraldehyde

Cited by 71 publications

References 23 publications

Integrated Micropillar Polydimethylsiloxane Accurate CRISPR Detection (IMPACT) System for Rapid Viral DNA Sensing

Integrated Micropillar Polydimethylsiloxane Accurate CRISPR Detection (IMPACT) System for Rapid Viral DNA Sensing

Effects of fixatives on myelin molecular order probed with RP-CARS microscopy

Expansion microscopy with conventional antibodies and fluorescent proteins

Contact Info

Product

Resources

About