What is an RFP (Request for proposal)?
An organization buying a service or a product issues the RFP document to request vendor bids for a product, solution or service and invites them to send their proposal that will meet the organization’s needs. Want to purchase services that will improve their business processes and bottom-line
What is the GFP gene?
Gfp refers to the gene that produces green fluorescent protein. Using DNA recombinant technology, scientists combine the Gfp gene to a another gene that produces a protein that they want to study, and then they insert the complex into a cell.
How do you use anti RFP reagent?
Simply add the reagent to your cells, incubate overnight, and you’re ready to image your cells. Anti-RFP antibodies provide a convenient method for visualizing RFP, especially when amplification of the fluorescent protein of interest is necessary to overcome a dim or degraded signal.
How can genomics improve the utility of RFP?
Genetic engineering has improved the utility of RFP by increasing the speed of fluorescent development and creating monomeric variants. Improved variants of RFP include mFruits (mCherry, mOrange, mRaspberry), mKO, TagRFP, mKate, mRuby, FusionRed, mScarlet and DsRed-Express.
Where does the RFP gene come from?
Recently, the majority of RFPs have been isolated and cloned from Anthozoa species living in the Indo-Pacific region (Piatkevich et al., 2010a; Verkhusha et al., 2003a).
What is the difference between RFP and GFP?
The RFP emission filter permits signals with 575 nm to 640 nm to pass through and the GFP emission filter permits fluorescent signals with wavelengths of 510 nm to 523 nm to pass through. These images confirm that the emission filter blocks unwanted signals efficiently and provide a clean image.
Is mCherry and RFP the same?
mCherry is a member of the mFruits family of monomeric red fluorescent proteins (mRFPs). As a RFP, mCherry was derived from DsRed of Discosoma sea anemones unlike green fluorescent proteins (GFPs) which are often derived from Aequoera victoria jellyfish.
What is RFP antibody?
Red fluorescent protein (RFP) is a biological marker used for monitoring physiological processes, visualizing protein localization, and detecting transgenic expression in vivo.
Is RFP derived from GFP?
Most fluorescent proteins were derived from GFP, but RFPs do vary in terms of their source. Perhaps RFPs derived from Discosoma sp would be useful since dsRed is structurally very similar.
Can you use GFP and RFP together?
Abstract. Red fluorescent proteins (RFPs) combined with GFP are attractive probes for double-fluorescence labeling of proteins in live cells.
Are DsRed and RFP the same?
DsRed is a red fluorescent protein (RFP) from coral Discosoma sp. [1], with the excitation and emission maxima at 558 and 583 nm, respectively [2]. DsRed is homologous to green fluorescent protein (GFP), which forms an 11-strand β barrel and a chromophore embedded inside the barrel [3].
Where is RFP purified?
Clusterin-RFP-GFP must be purified from mammalian cells, because clusterin is a secreted protein modified by glycosylation, disulfide bonds, and heterodimerization of two subunits.
What is the best 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.
Can RFP antibody detect mCherry?
This polyclonal RFP antibody detects RFP and its variants, such as mCherry, tdTomato, mBanana, mOrange and others. The RFP antibody has been validated for various applications and can be used for the detection of RFP and derivatives by ELISA, immunofluorescence, FACS, immunohistochemistry or western blotting.
Does RFP antibody recognize mCherry?
The anti-RFP antibody 5F8 recognizes common RFP variants including mCherry, mPlum, mRFP, mRFPruby.
What is RFP in business?
What is a request for proposal (RFP)? A request for proposal (RFP) is a document that an organization, often a government agency or large enterprise, posts to elicit a response -- a formal bid -- from potential vendors for a desired IT solution.
Does GFP antibody recognize RFP?
A: The anti-GFP antibody was made using the N-terminal of GFP as an antigen and will not react with RFP proteins dsRED or mCherry, since their sequences are so different.
Why can GFP be used in vivo?
Fluorescent proteins generally have very high extinction coefficients ranging up to approximately =95 000. In addition, fluorescent proteins have very high quantum yields up to 0.8. These properties make fluorescent proteins very bright. Two-photon absorption of GFP is important for deep-tissue imaging in vivo.
What is RFP in business?
What is a request for proposal (RFP)? A request for proposal (RFP) is a document that an organization, often a government agency or large enterprise, posts to elicit a response -- a formal bid -- from potential vendors for a desired IT solution.
How can I boost my GFP signal?
GFP fluorescence is detectable in sections post-fixed with PFA at 25 °C, although as a diffuse signal. By increasing the temperature of PFA to 30 °C or 37 °C, the GFP signal is stronger and more confined compared to brain sections post-fixed with PFA at 25 °C (arrows).
What is red fluorescent protein?
Red fluorescent protein ( RFP) is a fluorophore that fluoresces red-orange when excited. Several variants have been developed using directed mutagenesis. The original was isolated from Discosoma, and named DsRed. Others are now available that fluoresce orange, red, and far-red.
How long does it take for fluorescent proteins to mature?
Issues with fluorescent proteins include the length of time between protein synthesis and expression of fluorescence. DsRed has an maturation time of around 24 hours, which can make it unusable for many experiments that take place in a shorter time frame.
What are GFP markers?
Genetically encoded markers based on the GFP-like proteins have several advantages over the fluorescent dyes for applications in cell biology. FPs possess a unique ability to produce fluorescence after their expression in any prokaryotic or eukaryotic cell without additional enzymes or cofactors except for molecular oxygen. Moreover, a protein of interest can be easily tagged with an FP on a DNA level without affecting its intrinsic function. Consequently, FPs as reporter markers allow for studying temporal and spatial expression of genes by measuring their fluorescence signal in live cells and tissues, as well as localization and dynamics of cellular proteins, organelles, and virus particles. The recent expansion of spectrally distinct FP variants has enabled multicolor imaging for monitoring several events simultaneously.
What is FP technology?
Coupled with flow cytometry, the FP technology provides an amazing opportunity to noninvasively differentiate between various cell populations, monitor gene activities, and detect protein–protein interactions and small molecules in individual living cells in a high-throughput manner . Unfortunately, as yet RFPs have been underutilized in the flow cytometry and fluorescence-activated cell sorting (FACS) approaches. Recent advances in FP development and invention of the lasers emitting a variety of wavelengths greatly increase the performance and the capabilities of a multiparameter analysis with the FP-based flow cytometry. This chapter focuses on the novel improved RFPs, which are potentially useful for many FACS applications. We also describe the techniques for the utilization of RFPs as reporters and biosensors for the multiparameter flow cytometry.
Why is photostaability important in FPs?
Photostability of FPs can be important if postsort analysis by fluorescent microscopy is required . A comparison of bleaching half-times revealed that the DsRed, TagRFP-T, and mOrange2 proteins are the most photostable among the RFPs (Shaner et al., 2008). When choosing an FP for a particular experiment, it should be kept in mind that protein photostability strongly depends on the conditions of the protein biosynthesis and photobleaching (Bogdanov et al., 2009).
What are the steps of FP maturation?
FP maturation includes two consecutive steps: protein folding and chromophore formation, the latter usually being a rate-limiting step. The tertiary structure of all known FPs is highly conserved and organized as a capped β-barrel that serves as a shell to prevent chromophore quenching by solvent molecules (Day and Davidson, 2009). The main peculiarity of GFP-like fluorescent proteins is that the formation of the chromophore responsible for protein fluorescent properties occurs without any cofactors or enzymes but requires molecular oxygen. The chromophore is generated as a result of several consecutive autocatalytic reactions involving internal amino acid residues of the FP. Even though the first RFPs were cloned in 1999, it was not until recently that the actual mechanism of red chromophore formation has been revealed. It was shown that the posttranslational modifications of RFPs are more complicated than those for GFPs and do not always lead to the red chromophore formation (Piatkevich and Verkhusha, 2010).
What is the brightness of a FP?
Brightness of an FP is one of its most important characteristics. High brightness of an FP is usually an additional advantage for any cell biology application. Brightness determines the sensitivity and signal-to-noise ratio for the fluorescence detection. By definition, the molecular brightness of FP is the product of a molar extinction coefficient and quantum yield, measured in vitro. Since the extinction coefficient is a function of the wavelength, the effective brightness of an FP depends on the wavelenght of the excitation light. Moreover, the effective brightness is determined by emission filter sets used for detecting fluorescence (Fig. 1). Molecular brightness and effective brightness specified for certain laser excitations for common RFPs are presented in Tables Iand andII,II, respectively.
How to prevent FP fusion mislocalization?
The analysis of the amino acid sequence of an FP, particularly the amino acid residues external to the protein fold and the amino acid content of the N- and C-termini, can help to prevent FP fusion mislocalization. External cystein resiudes and N-glycosylation sites induce an incorrect folding and mislocalization of FP fusions due to their oligomerization in endoplasmic reticulum (Jain et al., 2001). Sometimes an FP can contain short localization sequences that disturb the original localization of a targeted protein (Kredel et al., 2009). Undesirable FP fusion behavior can be eliminated by a substitution of all external cysteins and glycosilation sites and by truncation of targeting domains.
What is the green fluorescent protein used for?
Chalfie et al.(1994)showed that a green fluorescent protein (GFP) from the jellyfish Aequorea victoriacould be used as a marker for protein localization and expression in living bacteria and worm cells. Cloning of GFP and it first application in vivodramaticly altered the nature and scope of the issues that could be addressed by cell biologists. Together with the introduction of new microscopy techniques, fluorescent proteins (FPs) changed the way life science research is performed today.
What is an RFP?
The RFP defines the project, for the company that issues it as well as the companies that respond to it. The RFP describes the project, its goals, and the organization that is sponsoring it and outlines the bidding process and contract terms. RFPs are used by most government agencies and many private companies and organizations.
What is an RFP in government?
In government, the RFP has been adopted as a way to ensure that cronyism is removed as a factor in the rewarding of contracts.
What Is a Request for Proposal (RFP)?
A request for proposal (RFP) is a business document that announces a project, describes it, and solicits bids from qualified contractors to complete it. Most organizations prefer to launch their projects using RFPs, and many governments always use them.
How does an RFP work?
The RFP process begins with drafting a request for proposal. Bidders review the solicitation and submit suggestions for improvement. After implementing feedback, the final request for proposal is issued. Bidders then submit their proposals.
Why do businesses use RFP?
In such cases, the RFP gives the business an efficient way to recruit the expertise they need to get the project done.
Why is RFP important?
In government, the RFP has been adopted as a way to ensure that cronyism is removed as a factor in the rewarding of contracts. It also opens up the process to competition, which can be expected to keep project costs lower.
Why do we need RFPs?
They are generally required to open up competition among private companies and remove bias from the process. The agencies want to ensure that they get the lowest and most competitive bid.
What is an RFP and When to Use it?
RFP is a formal document outlining a company’s intent and effort to secure a service, solution or a product.
What Should an RFP Document Include?
An RFP document is not a one-size-fits-all solution. It needs to be tailored to each business’s specific needs and according to the type of service or solution the company requires.
RFP Process: How to Write an RFP and Use It for Successful Procurement Procedure
Once you understand what elements constitute an average request for proposal document, it’s time to start drafting it.
RFP Software Solutions
Plenty of business organizations use software solutions to streamline the RFP process since these documents involve a lot of moving parts and can be quite time-consuming to make.
The Benefits of RFP
Issuing an RFP helps businesses find the best vendor for their needs and minimizes risk of poor ROI.
Software Development RFP
RFP for software development is the initial document that you can use when looking to select a software development partner, specifically.
Mobile App Development RFP
Similar to software development RFPs, a mobile app development RFP is a procurement document companies draft and send to vendors when they need to have a mobile app developed.
What is the shape of a GFP protein?from blog.addgene.org
The protein structure, first reported in 1996, is an eleven β-sheet-containing “barrel” shape , with the chromophore concealed at the center of the structure, shielded from quenching by aqueous solvent. This tightly-packed structure explains the importance of the entire GFP protein, which is almost completely required to maintain fluorescent ...
Why do plasmids have GFP?from blog.addgene.org
Cell marking/selection: Expression constructs like plasmids often include GFP as a marker to help identify which cells have successfully taken up the plasmid. This can serve as an alternative to antibiotic selection. Plasmids of this type may have the GFP under the control of an additional promoter from that of the gene of interest, or expressed from the same transcript as the gene of interest, but after an internal ribosome entry site (IRES). This is oftentimes used in conjuction with FACS (see below).
Why Green Fluorescent Protein?from blog.addgene.org
It has a fluorescent emission wavelength in the green portion of the visible spectrum (hence the name), which is due to a chromophore formed from a maturation reaction of three specific amino acids at the center of the protein (Ser65, Tyr66, and Gly67). When first discovered, one of the most surprising aspects of GFP was the fact that the chromophore forms spontaneously and without additional co-factors, substrates, or enzymatic activity – it only requires the presence of oxygen during maturation. This meant that the protein could be taken directly from A. Victoria and expressed in any organism while still maintaining fluorescence.
What is a GFP biosensor?from blog.addgene.org
Biosensors: A wide array of GFP-based fluorescent biosensors has been designed to detect a variety of intracellular conditions, including ion (such as Ca2+) concentrations and pH, using a range of strategies such as FRET, calmodulin, and others. Review Addgene's collection of fluorescent biosensors here.
What is FRET in biology?from blog.addgene.org
Förster resonance energy transfer (FRET): This is used to study the interactions between two proteins or between two domains of a protein that undergoes conformational change. Typcially two fluorescent proteins with overlapping excitation/emission spectra are used; one fused to each protein or domain being tested. Find FRET plasmids here.
Why is GFP used in cell fate studies?from blog.addgene.org
Developmental/transgenic uses: Because of its stability, GFP can be used in lineage tracking capacities in cell fate studies. It can also be used, when put under control of promoters of interest, to visualize the developmental stage at which these promoters are active.
Why did scientists develop new versions of GFP?from blog.addgene.org
Almost as soon as its sequence was elucidated, scientists began engineering new versions of GFP through mutagenesis in order to improve its physical and biochemical properties. In 1995, Roger Y. Tsien described an S65T point mutation that increased the fluorescence intensity and photostability of GFP. This also shifted its major excitation peak ...
Who discovered the GFP gene?
In the early 1990s, molecular biologist Douglas Prasher, at the Marine Biology Laboratory, used GFP to design probes, a technology involving fragments of DNA to detect the presence of nucleotide sequences. Prasher isolated the complementary DNA (cDNA) of Gfp gene, and he published the sequence of the gene in 1992.
How do scientists combine the GFP gene?
Using DNA recombinant technology, scientists combine the Gfp gene to a another gene that produces a protein that they want to study, and then they insert the complex into a cell. If the cell produces the green fluorescence, scientists infer that the cell expresses the target gene as well.
What protein did Chalfie discover?
Chalfie heard about the protein GFP in a lecture, and he speculated that GFP might facilitate his study of gene expression in C. elegans. Chalfie's team obtained the cDNA of the gene Gfp from Prasher and inserted only the coding sequence of Gfp gene first in the bacterium Escherichia Coli, and then in C. elegans.
How does GFP affect biology?
As GFP does not interfere with biological process es when used in vivo, biologists use it to study how organisms develop. For example, after 1994, Chalfie and his colleagues applied GFP in the study of the neuron development of C. elegans. In a 2002 paper, Chalfie and his colleagues describe how they first labeled a specific gene involved in tactile perception in neuron cells with GFP, and then observed the amount of fluorescence emitted by those cells. Because mutant cells produced less or more GFP than normal cells, the abnormal amount of fluorescence production indicated the abnormal development of mutants. Since then, this field of research expanded to many other organisms, including fruitflies, mice, and zebra fish.
What is the green fluorescent protein?
Green Fluorescent Protein. Green fluorescent protein (GFP) is a protein in the jellyfish Aequorea Victoria that exhibits green fluorescence when exposed to light. The protein has 238 amino acids, three of them (Numbers 65 to 67) form a structure that emits visible green fluorescent light. In the jellyfish, GFP interacts with another protein, ...
What is the name of the protein that is used to study cells in embryos and fetuses?
The protein has 238 amino acids, three of them (Numbers 65 to 67) form a structure that emits visible green fluorescent light. In the jellyfish, GFP interacts with another protein, called aequorin, which emits blue light when added with calcium. Biologists use GFP to study cells in embryos and fetuses during developmental processes.
Why do scientists use GFP?
Biologists use GFP to study cells in embryos and fetuses during developmental processes. Biologists use GFP as a marker protein. GFP can attach to and mark another protein with fluorescence, enabling scientists to see the presence of the particular protein in an organic structure. Gfp refers to the gene that produces green fluorescent protein.
What Is A Request For Proposal (RFP)?
Red fluorescent protein (RFP) is a fluorophore that fluoresces red-orange when excited. Several variants have been developed using directed mutagenesis. The original was isolated from Discosoma, and named DsRed. Others are now available that fluoresce orange, red, and far-red.
RFP is approximately 25.9 kDa. The excitation maximum is 558 nm, and the emission maximum is 583 nm.
Understanding A Request For Proposal
Requirements For A RFP
Benefits of A RFP
- RFPs are used for complex projects, often requiring a number of sub-contractors. They describe the organization issuing the RFP, the scope of the project being undertaken, and the criteria for evaluating entries. They also outline the bidding process and the contract terms. The requests include a statement of work describing the tasks to be performed by the winning bidder and the …
RFP vs. RFQ vs. RFI
- Government agencies or other entities may be required to issue requests for proposals to provide full and open competition and to drive down the cost of a solution. Accepting a proposal that is most responsive to specifications may not always mean the lowest-priced bid. Skillfully creating a request for proposal can ensure the success or failure of the resulting solution. If the …
Example of A RFP
- An RFP is, in part, an advertisement. It announces that a project is proceeding, and opens the door to qualified candidates who can get the job done. In government, the RFP has been adopted as a way to ensure that cronyism is removed as a factor in the rewarding of contracts. It also opens up the process to competition, which can be expected to keep project costs lower. The alternative t…
The Bottom Line
- A request for proposal (RFP), request for quote (RFQ), and request for information (RFI) are three distinct types of documents that businesses and other entities use to reach out to the business community for suppliers or contractors that they may be able to work with. 1. A request for proposal, as noted above, announces a specific project that is planned and solicits contractors c…