A guide to choosing fluorescent proteins (2024)

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A guide to choosing fluorescent proteins (2024)

FAQs

Which fluorescent protein should I use? ›

For single-color experiments, green FPs are the most common choice. EGFP is the most popular green FP and is a good choice for many single-color studies. However, other green FPs, such as TurboGFP (a.k.a. maxGFP), may be better choices for certain applications.

Is DsRed the same as RFP? ›

While DsRed is a specific type of RFP, the term RFP encompasses a wider range of red fluorescent proteins, including DsRed. Advances in genetic engineering have led to the development of numerous RFP variants with improved properties for various research and industrial applications.

What are the most commonly used fluorescent proteins? ›

Green fluorescent protein (GFP) from jellyfish Aequorea victoria is the most extensively studied and widely used in cell biology protein. GFP-like proteins constitute a fast growing family as several naturally occurring GFP-like proteins have been discovered and enhanced mutants of Aequorea GFP have been created.

What is the difference between mCherry and RFP? ›

mCherry is a red fluorescent protein (RFP), classified as a luminescent protein, that belongs to a group of fluorescent protein chromophores. mCherry is a part of the mFruits protein family, which is a family of mRFPs, monomeric red fluorescent proteins.

How do I choose fluorochromes? ›

Choose the brightest fluorochrome for your least expressed protein and the dimmest fluoro- chrome for your most highly expressed protein. (View our Expression of Common Cell Surface Proteins chart to assist you.) Generally, on leukocytes, CD45 is one of the most highly ex- pressed cell surface proteins.

Is YFP better than GFP? ›

GFP emits a green fluorescence when exposed to blue ultraviolet light. The improved versions of YFP have reduced chloride sensitivity and faster maturation. They also have increased brightness due to quantum yield.

What is the most stable red fluorescent protein? ›

mCherry is the most widely used and cited red fluorescent protein owing to its fast maturity, stability, and resistance to photobleaching. mCherry and DsRed-Monomer are ideal for tagging proteins with diverse functions and/or subcellular localization patterns.

What is the difference between red fluorescent protein and green fluorescent protein? ›

Fluorescent proteins (FPs) form a fluorophore through autocatalysis from three consecutive amino acid residues within a polypeptide chain. The two major groups, green FPs (GFPs) and red FPs (RFPs), have distinct fluorophore structures; RFPs have an extended π-conjugation system with an additional double bond.

Why does RFP fluoresce red? ›

The red fluorescence displayed by these proteins arises from the presence of an acylimine group conjugated with the standard p-hydroxybenzylideneimidazolinone GFP chromophore (6). The additional double bond extends the size of the chromophore conjugated system leading to an increase in emission wavelength.

Why use green fluorescent protein? ›

GFP makes for an excellent tool in many forms of biology due to its ability to form an internal chromophore without requiring any accessory cofactors, gene products, or enzymes / substrates other than molecular oxygen. In cell and molecular biology, the GFP gene is frequently used as a reporter of expression.

What is the smallest brightest fluorescent protein? ›

With molecular weight of only 17 kDa, miRFP670nano is the smallest monomeric NIR FP that fluoresces in mammalian cells as bright as twice bigger state-of-art two-domain NIR FPs.

What is the difference between GFP and enhanced green fluorescent protein? ›

The difference between the fluorescence of the green fluorescent protein (GFP) and the enhanced green fluorescent protein (EGFP) is that EGFP emits 35× the fluorescence of GFP when excited with ultraviolet or blue light: it is much brighter.

What are the disadvantages of mCherry? ›

mCherry is a very popular red fluorescent protein. However it has several disadvantages: It is shifted towards far red (ex peak 585 nm) so it often is not imaged optimally with the illumination sources and filters commonly available. It bleaches fast.

Can you use GFP and RFP together? ›

Red fluorescent proteins (RFPs) combined with GFP are attractive probes for double-fluorescence labeling of proteins in live cells.

Can you use GFP and mCherry together? ›

Due to the spectral properties of GFP and mCherry, they are considered an ideal combination for co-localisation and co-expression experiments.

What is the difference between Venus and EGFP? ›

The Venus protein differs from EGFP and EYFP by 8 and 5 amino acid exchang- es, respectively. It has the same spectral properties as EYFP, but it has a 30-fold higher relative fluorescence, it matures faster and is less sensitive to pH and chloride ion concentrations (7).

Is neon green better than GFP? ›

The monomeric GFP mNeonGreen is about 3- to 5-times brighter than GFP and monomeric enhanced GFP and shows high photostability. The maturation half-time of mNeonGreen is about 3-fold faster than that of monomeric enhanced GFP.

What is the difference between EGFP and mGFP? ›

Compared to EGFP, mGFP provides about the same brightness of fluorescence but is significantly more pH stable. mGFP is specially optimized for expression at 37°C.

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