Protein expression, isolation and purification experiments
Protein expression, isolation and purification can: (1) explore and study the function of genes and the mechanism of gene expression regulation; (2) be used for
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用于检测、定量和定位抗原与抗体的技术——ELISA、免疫印迹、免疫组化与流式细胞术。以下是我们知识库中标注该主题的实验方案、常见问题与技术文章。
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Protein purification experiments
Protein molecules of different sizes enter the colloidal filtration column and can be separated according to their molecular weight differences; it is a widely
Protein extraction experiments
Protein extraction experiments are basic operations for performing experimental analysis of proteins and can be used for (1) protein isolation (2) protein funct
Preparation of protein gel electrophoresis gels
Protein gel electrophoresis is commonly used to (1) analyze molecular biology, genetics, and biochemistry (2) preparative techniques (3) partially purify molecu
Immunoglobulin extraction technology
The immunoglobulin extraction technique can be applied to (1) infer the body's humoral immune function, and (2) diagnose high and low Ig caused by certain disea
protein sequencing
Protein sequencing can be used to: (1) identify proteins; (2) characterize post-translational modifications of proteins. (3) Analyze the relationship between pr
Measurement of plasma free hemoglobin
Measurement of plasma free hemoglobin can be used to (1) approximate the value of red blood cells used. (2) Elevations and decreases in hemoglobin can be refere
Serum Protein Agarose Gel Electrophoresis
Serum protein agarose gel electrophoresis can be applied to (1) determine the protein content of serum and (2) diagnose diseases.
Serum Preparation Experiment
Antisera are sera containing a certain class of antibody molecules with specific immune functions, generally prepared from animals that have been artificially i
vise
The study of membrane ion channel properties using the membrane clamp is an arduous, detailed and complicated work, which requires a high level of technical lev
Nuclease protection assay
Ribonuclease protection assay (RPA) is a new method for quantitative analysis of mRNA developed in the last decade. The basic principle is to hybridize the labe
Solid phase synthesis, cleavage and purification of peptides
Solid-phase peptide synthesis technology is a key technology of modern protein chemistry, and also a technology of great influence and significance to modern mo
High Performance Resistance Liquid Chromatography
High performance resistive liquid chromatography, also known as volumetric resistive chromatography (SEC), molecular sieve chromatography, gel filtration, etc.
High performance reversed-phase liquid chromatography
High performance reversed-phase liquid chromatography can be applied to: Separate non-polar, polar or ionic compounds.
ion exchange chromatography
Ion exchange chromatography is a chromatographic method in which ion exchange genes (CM, SP, Q, DEAE, etc.) are bonded to certain inert carriers (cellulose, cro
Protein extraction and crude separation experiments
To isolate and purify proteins, the target protein is firstly extracted from the raw material, then a large number of impurities are removed by the method of co
Determination of urease activity in soybean meal
Crushed soybean products were mixed with a neutral urea buffer solution and kept at 30°C for 30 min before urease catalyzed the reaction of urea hydrolysis to
Determination of tissue level metallothionein by cadmium-hemoglobin method
Metallothionein (MT) is a metal- and sulfur-rich inducible protein, which can be induced by certain metals (Bi, Zn, Cu, etc.), hormones (glucocorticoids), cytot
Determination of glutathione-S-transferase activity
Glutathione S-transferases are enzymes that catalyze the reaction of exogenous compounds with electrophilic substituents with endogenous reduced glutathione (GS
Determination of glucuronide-transferase activity
Glucuronosyltransferases are enzymes that catalyze the combination of indirect bilirubin with glucuronic acid to form glucuronides (i.e., direct bilirubin).
Determination of aniline-4-hydroxylase activity
Hydroxylase, also known as hydroxylase, is a type of oxygenase, an enzyme that catalyzes the reaction that utilizes oxygen molecules to form hydroxides (phenols
Induction of heparanase
Hepatic drug enzymes is an abbreviation for the mixed function enzyme system of liver microsomes. This enzyme system is a group of mixed-function oxidative enzy
Determination of sodium, potassium-triphosphate adenosine triphosphatase activity
ATPases, also known as adenosine triphosphatases, are a class of enzymes that catalyze the hydrolysis of adenosine triphosphate (ATP) to adenosine diphosphate (
Extraction of myocardial sarcoplasmic reticulum calcium-triphosphate adenosine monophosphatase
Cardiac sarcoplasmic reticulum calcium-ATPase (SERCA2a) plays a major role in the regulation of calcium homeostasis in cardiomyocytes, and studies on the gene t
Determination of the activity of myocardial sarcoplasmic reticulum calcium-triphosphate adenosine monophosphatase
Cardiac sarcoplasmic reticulum calcium-ATPase (SERCA2a) plays a major role in the regulation of calcium homeostasis in cardiomyocytes, and studies on the gene t
Metallothionein Measurement Experiment
Metallothionein is a short cysteine-rich peptide produced by microorganisms and plants as a metal-binding protein with a high affinity for a wide range of heavy
Immunofluorescence localization experiments of nuclear proteins
Immunofluorescence localization of nuclear proteins can be applied to (1) localize intracellular distribution of nuclear proteins and (2) viral studies.
Immunoscreening assay
A technique for selecting a target from a population based on the principle of antigen-antibody interaction. For example, antibodies are used to detect proteins
Protein recovery assay
Recovery test is a type of "control test". When the analyzed sample is complex and not completely clear, a known amount of the measured component is added to th
affinity chromatography
The affinity molecules with special structure are made into solid-phase adsorbent and placed in the chromatography column, when the protein mixture to be separa
Molecular exclusion high performance liquid chromatography experiments
Molecular exclusion chromatography, also known as gel chromatography, is the use of different molecular sizes of the separated substances lead to different degr
Characterization of decoy proteins
The first step in capturing interacting proteins is to construct a decoy protein, which is a fusion of LexA to the target protein. A series of control experimen
Capture experiments of interacting proteins
Capture assays for interacting proteins are subjected to two sequential bulk yeast plate screening processes. Saccharomyces cerevisiae contains the lexA fusion
Probe synthesis experiments
A probe is a small fragment of single-stranded DNA or RNA (approximately 20 to 500 bp) that is used to detect a nucleic acid sequence complementary to it. Doubl
Expression and purification experiments of glutathione S-transferase fusion protein
The pGEX vectors can be used to express and purify peptides (including short peptides) used as immunogens and biochemical and biological reagents, or to constru
Expression and purification experiments of thioredoxin fusion protein
A gene fusion expression system using the E. coli trxA gene product-thioredoxin as a fusion partner is particularly suitable for the expression of high yield so
Metabolic labeling experiments of recombinant proteins
Since host protein synthesis is essentially terminated when recombinant proteins are expressed, in vivo metabolic labeling is a sensitive method for detecting r
Experiments to determine peak protein production
Protein is the material basis of life, without which there is no life. It is therefore a substance that is closely associated with life and with all forms of vi
Phosphate amino acid analysis experiment
It is of interest to identify phosphorylated amino acid residues in proteins. Phosphorylation occurs at serine, threonine, and methionine in proteins, and the i
Electrotransfer experiments on proteins
Many membranes can be used as solid-phase supports for protein electrotransfer, such as diazotized cellulose membranes (DPT, DBM), DEAE-cellulose membranes, nyl
Salt chromatographic graded separation and gel chromatographic desalting of proteins
The purpose of this experiment is to understand the basic principles and operation of salting out graded separation of proteins; to understand the basic princip
Molecular weight determination of proteins by gel filtration chromatography experiment
Gel filtration is also known as exclusion chromatography, molecular sieve chromatography and gel chromatography. The purpose of this experiment is to understand
Electrophoretic separation and identification of proteins on cellulose acetate membranes
The isoelectric points of various proteins in mixed protein samples are different, and the amount of electrostatic charge carried by various proteins is differe
Analysis of N-terminal amino acids of proteins by dansulfonylation method
The reaction of α-amino group of protein with dansulfonyl chloride (DNS-Cl is a fluorescent substance) produces DNS-protein, which can be hydrolyzed to produce
Separation of nucleotides by chromatography on ion exchange columns
In this experiment, yeast RNA was used as the material, and the RNA was hydrolyzed into mononucleotide by alkali, then separated by ion exchange column chromato
Experiments on the preparation of yeast sucrase
Sucrase (inver tase) (β-D-fructofuranoside fructohydrolase) (EC. 3 . 2 . 1 . 26 ) catalyzes the hydrolysis of α-fructofuranoside bonds in non-reducing sugars
The effect of substrate concentration on catalytic reaction rate and the determination of Mie constant Km and maximum reaction velocity Vmax
In this experiment, the Km value of sucrase was calculated by using the amount of products (glucose and fructose) formed by sucrase hydrolyzing different concen
Protein sequence analysis and structure prediction experiments
Source of content: Guangdong Pharmaceutical University Laboratory Instruction Manual.
Protein ultraviolet spectrometry assay
Since the benzene rings of tyrosine and tryptophan residues in protein molecules contain conjugated double bonds, proteins have the property of absorbing ultrav
Serum lipoprotein agarose electrophoresis assay
After serum lipoproteins are stained with Sudan black B and electrophoresed in pH 8.6 barbiturate buffer with agarose, the lipoproteins can be divided into diff
Polyacrylamide gel electrophoresis of serum proteins
Serum protein polyacrylamide gel has the advantages of good mechanical strength, elasticity, transparency, high chemical stability, non-electroosmosis, simple e
Titration curve experiment
The titration curve is a curve presenting the mobility of each component of the sample against pH, for which the gel containing the carrier amphoteric electroly
Activation assay of calmodulin samples
This experiment describes the process of calreticulin sample activation measurement. This experiment is from the Experimental Guide for Protein Purification and
Experimental extraction of calmodulin from chicken gizzard
Calmodulin is found in all mammalian tissues and also in other eukaryotic organisms. It is one of the most ubiquitous and highly conserved proteins known. This
Crude graded separation experiments of calmodulin
For the purification of calmodulin, two different operations were performed for the crude graded separation of smooth muscle tissue extracts: ammonium sulfate p
DEAE SEPHADEX A-50 anion-exchange chromatography assay for calmodulin
The next step in the purification of calmodulin is anion exchange chromatography with DEAE resin. This step takes advantage of the fact that calmodulin is a str
Phenyl-Sepharose chromatographic assay of calmodulin
This protocol has been developed over many years of practice, and it is not an example of a first attempt to purify a protein by HIC chromatography. For calmodu
Urea gradient polyacrylamide gel electrophoresis and folding analysis experiments
Goldenberg and Creighfn (1984) introduced the use of polyacrylamide gels containing a concentration gradient of urea to analyze the folded state of proteins. A
Cyanogen bromide cleavage assay of calmodulin
This experiment describes the cyanogen bromide cleavage process of calmodulin. This experiment is from the Experimental Guide for Protein Purification and Ident
Trypsin digestion assay of calmodulin
This experiment describes the trypsin digestion process of calmodulin. This experiment was derived from Protein Purification and Identification Laboratory Guide
Experiments on the preparation of nuclear extracts from HeLa cells
In this experiment, nuclear extracts were prepared from HeLa cells essentially as described by Dignam et al. (1983). This basic method is probably the most comm
How to load gel filtration column experiment?
This experiment describes how to load a gel filtration column. This experiment is from Protein Purification and Identification Laboratory Guide by Houzhu Zhu.
Sephacryl S-300 HR column chromatography of HeLa cell nucleus extracts
This experiment describes the process of Sephacryl S-300 HR column chromatography of HeLa cell nucleus extract. This experiment is from the guide to protein pur
Experiments on purification of oligonucleotides by preparative gel electrophoresis
This experiment describes how to purify oligonucleotides by preparative gel electrophoresis. This experiment is from Protein Purification and Identification Lab
Preparation of DNA affinity media
This experiment describes the preparation process of DNA affinity medium. This experiment is from the Laboratory Guide for Protein Purification and Characteriza
Operational experiments in affinity chromatography
This experiment introduces the related operation of affinity chromatography. This experiment is from the Laboratory Guide for Protein Purification and Identific
Gel mobility change analysis experiment
In this experiment, we used the same complementary oligonucleotide used for DNA affinity chromatography (see the end of the Introduction to Experiment 3 for the
Heparin-Sepharose CL-2B preparation experiment
This experiment describes the preparation of heparin-Sepharose CL-2B. This experiment was derived from the Experimental Guide for Protein Purification and Ident
Experiments on the fragmentation of Escherichia coli cells and preparation of inclusion bodies
This experiment describes the fragmentation of E. coli cells and the preparation of inclusion bodies. This experiment is from the Laboratory Guide for Protein P
immunoaffinity chromatography
This experiment introduces the method of immunoaffinity chromatography. This experiment is from the Laboratory Guide for Protein Purification and Identification
Quantitative measurement of proteins
There is a great need for a rapid and accurate method for quantitatively determining the protein concentration in a sample so that one can follow the recovery o
Quantitative protein spot blotting assay
Protein blotting, which uses immunological methods to measure the amount of a particular protein. This experiment was derived from Protein Purification and Iden
Enzyme assay experiment
In this unit, a simpler method was used to determine its stimulatory effect on core RNA polymerase transcription from a non-specific template poly(dAT) start. T
Purity estimation experiment by overloaded stained SDS gel scanning method
Since proteins can be separated with high resolution on SDS gels and sensitive staining can be given with Caumas Brilliant Blue (CBB), the purity of the protein
Evaluation of homogeneity experiments by nondenaturing gel electrophoresis mobility shift rate method
When a protein is electrophoresed under conditions that keep its natural conformation constant, its mobility can be used as a measure of its homogeneity. Since
Rapid protein spot blotting assay
In routine protein spot blotting, the blotting membrane is closed for 1 hour, reacted with primary antibody for 1 hour, and then reacted with secondary antibody
Inclusion body precipitation dissolution assay
This experiment describes the amount of sodium N-dodecyl sarcosinate required for solubilizing inclusion body precipitation. This experiment is from the Laborat
Precipitation σ32 and RNA polymerase assays
This experiment describes the amount of polyethyleneimine needed to precipitate σ32 and RNA polymerase. This experiment was derived from Protein Purification a
Protein binding assay with immunoaffinity mediators
This experiment describes how much time it takes for a protein to bind to an immunoaffinity medium. This experiment is from the Protein Purification and Identif
New purification experiments of bacterial overexpression products (without disulfide bonds)
The purpose of this section is to provide an abbreviated set of working procedures for how to use a series of experiments similar to those described in this uni
Neville's method for the separation of liver plasma membrane
This experiment describes the Neville method for isolation of liver plasma membrane. This experiment is from the guide to protein purification and characterizat
An alternative experimental approach to the isolation of crude plasma membrane fractions from tissues
The method described by Neville (1968) works well for isolating murine liver plasma membranes, but is not necessarily applicable for isolating plasma membranes
Measurement of protein concentration in the presence of interfering substances
Protein concentrations have traditionally been determined by the method of Lowry and co-workers (Lowry et al., 1951) using the Folin-Ciocalteau reagent, but thi
Experiments for the determination of the number of insulin receptors on membranes
Insulin binds to its receptor with high affinity (KD~109 mol/L), and this ability allows us to estimate the number of insulin receptors in even fairly crude pre
Insulin receptor solubilization and activity assay experiments
This experiment describes about the process of solubilization and activity determination of insulin receptor. This experiment is from the guide to protein purif
Screening experiments of descaling agents for solubilizing membrane proteins
Although TriumX-100 is a good descaling agent for solubilizing insulin receptors from murine liver plasma membranes, it may not be suitable for solubilization o
Lectin affinity chromatography experiments on solubilized receptors
This experiment describes the process of lectin affinity chromatography on solubilized receptors. This experiment was derived from Protein Purification and Iden
Insulin affinity chromatography assay
In this experiment, we used insulin agarose column chromatography as the final step in insulin receptor purification. This method utilizes the high affinity bin
Coupling experiments of ligands with activation media
Here, we introduce this procedure by taking the coupling of ligand to Affi-Gel10 as an example. This experiment was derived from Protein Purification and Identi
Cross-linking experiments of the insulin receptor with [125I]insulin
This experiment describes about the cross-linking process of insulin receptor with [125I] insulin. This experiment is from the Experimental Guide for Protein Pu
Autophosphorylation experiments on insulin receptor stimulated by insulin
In this experiment, we will examine the insulin-stimulated tyrosine kinase activity of the insulin receptor. The phosphorylated receptor will be detected with a
Analytical experiments on glycosylation of insulin receptors
This experiment describes about the analysis of insulin receptor glycosylation. This experiment is from the Protein Purification and Identification Lab Guide by
silver dyeing experiments
This experiment describes the process of gel staining silver staining. This experiment is from the Protein Purification and Identification Laboratory Guide by Z
Precipitation experiment with trichloroacetic acid
TCA precipitation experiments are mainly used for (1) precipitation purification of proteins and (2) extraction of specimen compounds.
Acidified acetone/methanol precipitation method experiment
Another useful alternative to TCA precipitation is the use of an acidified acetone-methanol mixture. Acidification helps to remove SDS from the sample by dissol
Performic acid oxidation experiments on proteins
Oxidation of proteins with peroxyformic acid is a simple and effective method of partially denaturing proteins and stabilizing sulfur-containing amino acids. Pe
Separation of phosphopeptides with iminodiacetic acid (iron chelate) Sepharose 6B
In the structural and functional analysis of proteins (phosphorylated proteins), it is useful to be able to separate phosphopeptides from unmodified (unphosphor
Peptide isolation and purification experiments for protein internal sequence analysis
One of the most important aspects of molecular biology is the analysis of trace proteins, making possible the cloning of cDNA sequences encoding the proteins to
SDS-PAGE protein sample preparation
This experiment describes the preparation of SDS-PAGE protein samples.
Determination of isoelectric point of proteins by isoelectric focusing electrophoresis
Isoelectric focusing (IEF for short) is a new technique that appeared in the mid-sixties. In recent years, IEF has made new progress and has rapidly developed i
Experiments on combinatorial protein design strategies using computational methods
Computational methods have always played an important role in protein design. This work focuses on searching the protein sequence space to find one or several p
Integral adulteration experiments of non-natural amino acids in Escherichia coli
The doping of amino acid analogs is becoming increasingly useful. The targeted incorporation of non-natural amino acids has made it possible to use chemical bio
Experiments on design and optimization techniques for coiled spiral structures
Despite their apparent simplicity, coiled coil (Coiled Coil) modules are highly specialized and are important in understanding tertiary structure and its format
Calcium indicator experiments based on the fusion of calmodulin and fluorescent protein
Calmodulin (CaM) is a ubiquitously distributed protein involved in calcium-involved signaling. Upon Ca2+ influx, CaM acquires a strong affinity for binding vari
Design and synthesis experiments of artificial zinc finger proteins
Among DNA-binding modulators, (Cys)2(His)2-type zinc-finger modulators have a large potential for manipulation. Zinc finger modulators provide an attractive fra
Antibody simulation experiments based on the framework of fibronectin type III structural domains
We developed the use of the 10th fibronectin type III structural domain (FNfnlO) of human fibronectin as a framework to show the use of multiple surface chain s
Site-specific nucleic acid endonucleases for protein engineering experiments
Site-specific nucleic acid endonucleases are involved in many aspects of nucleic acid biochemistry. Restriction enzymes and related enzymes have become the para
Design and screening of protein libraries - a probabilistic calculation experiment
When designing protein libraries for screening, we must do everything possible to generate a diversity of protein variants within the physical limits of what ca
DNA fragmentation and directed evolution experiments using nucleotide exchange and shearing techniques
The use of DNA mashups to mimic natural evolutionary processes is a common approach to optimize DNA and protein properties. Here we present a new development in
Simplified oligonucleotide gene mixing assay
Improvements in the biochemical properties of proteases can be achieved through misdirected mutagenesis and DNA hybridization of the enzyme genes. The hybridiza
M13 Experiments on the application of phage coat protein modification in improved phage display technology
This chapter describes a method for improving the level of fusion protein presentation on the surface of M13 phage particles. Introduction of a mutation to the
Ribosome Inactivation Demonstration System Experiment
Here we present a new strategy for linking genotype and expression in vitro for the selection of functional proteins. In this strategy, the A chain of the ricin
Experiments in compartmentalized self-replication (a new approach to the directed evolution of polymerases and other enzymes)
Compartmentalized self-replication (CSR) is a novel approach to the directed evolution of proteases, especially polymerases. In the simplest form of CSR, it con
Synthesis of a parsimonious evolutionary library of the Ras-binding structural domain of the Raf protein and cloning experiments for rapid screening of dihydrofolate reductase using the fragment complementation method
The field of protein engineering is primarily concerned with the most fundamental sequence determinants regarding the design of new enzyme activities or folds,
Experiments on methods of applying end truncation, evolution, and re-elongation techniques to improve enzyme stability
Improving enzyme stability is an ideal design step for modifying enzymes to exercise their activity under extreme conditions, such as high temperatures, as well
Proteomic analysis of AtT20 pituitary cells as a model for neuroendocrine peptidergic system interference
Environmental contaminants can affect many intracellular enzyme activities. The effects of enzyme inhibitors on the proteome can be determined using bidirection
A proteomic approach to the risk assessment of peroxisome proliferating contaminants in the marine environment
Pollution of the marine environment is a growing concern because it can affect humans and other natural species. The application of proteomic methods to monitor
Maintenance of protein stability
This method is mainly used to maintain the stability of proteins.
Protein concentration and solute removal experiments
Outstanding growth is expected in novel primary molecules and biosimilar entities. Some of the advances are improved analysis, development and interactions. Num
Bacterial expression systems for exogenous protein production
Bacterial expression systems are available in a wide variety of vectors and host bacteria, and the short proliferation time of most engineered bacteria not only
Selection of suitable methods for recombinant protein expression
Recombinant proteins are important tools for studying biological processes. Expression systems are needed for their preparation. The choice of a suitable expres
Arbovirus-Insect Cell Expression System
This chapter will begin with an introduction to the basic baculovirus-insect expression system, Beijing Information, and then focus on recent advances that have
Experiments on the preparation of recombinant proteins by transient gene transfer in mammalian cells
Reports show that in the past decade, a variety of highly efficient transient transfection methods have been developed that meet or exceed these requirements. C
Refolding experiments after inclusion body proteolysis
When proteins are expressed at high levels in E. coli and other expression hosts, it is common to find that most of them are insoluble and exist in insoluble fo
Labeling experiments for protein expression
Most proteins are prepared using recombinant expression techniques. During the cloning process, additional residues or tags can be added to the N- or C-terminus
Experiments on the preparation of biological extracts required for protein purification
Proteomics and structural genomics require protein extraction methods that allow the simultaneous screening of hundreds of proteins on different combinations of
Gel filtration experiment
Unique among chromatographic techniques for protein purification, gel filtration separates proteins based on the relative size of their molecular masses. In con
Protein precipitation technology
Although many different precipitation methods have been used over the past 100 years, AS precipitation has been the most widely used, especially for acidic prot
Hydroxyapatite columns for protein chromatography experiments
Hydroxyapatite (HA) is a calcium phosphate-based hydroxylate that has been used extensively in the chromatographic separation of proteins, mainly in 1991-2009,
Protein gel staining assay
This chapter describes electrophoretic-based protein assays, focusing on the current methods for total protein assay, post-translational modification (PTM) assa
Identification, production and use of polyhydroxy-reactive monoclonal antibodies - for immunoaffinity chromatography
All forms of affinity chromatography require the formation of a specific interaction between two components by which one of the components can be purified. Immu
Purification experiments of membrane proteins
Membrane proteins play a key role in biological processes. They are involved in cell-cell and cell-matrix interactions, cell organelle and cell formation, trans
Protein elution experiments in gels
SDS-PAGE has proven to be an extremely useful analytical method in determining the number and size of peptides in a sample. It also has the ability to separate
Protein purity assay
No method can directly quantify the purity of a protein sample. Typically, protein purity determination involves evaluating the level of specific impurities or
Isoelectric spot focusing for protein separation
Proteins can be separated on coated or uncoated capillary columns, and the choice of separation protocol depends on the specific properties of the target protei
Analyzing peptide separation experiments
CE is particularly useful for separating specific peptides from complex mixtures such as proteolytic products. The following protocol describes the use of the P
Determination of phosphorylated amino acid positions in peptides
If the sample volume is insufficient for direct sequencing, the peptide can be degraded by manual Edman degradation to determine the location of the phosphoryla
Detection of specific amino acids in phosphopeptides by secondary discriminatory digestion assay
More information about phosphopeptides can be obtained by digestion with sequence-specific proteases or cleavage with site-specific chemicals. From The Compact
Preparation of monoclonal antibodies against phosphopeptides.
This experiment provides steps starting from inoculation of candidate hybridoma clones into a 96-well culture plate all the way through to isolation of a hybrid
Cyclic nucleotide-dependent protein kinase analysis assay
Protein kinases are analyzed using labeled donor substrates, and the accumulation of markers in protein or peptide acceptor substrates is readily detected when
Ca2+/calmodulin-dependent kinase analysis assay
Protein kinases are analyzed using labeled donor substrates, and the accumulation of markers in protein or peptide acceptor substrates is readily detected when
Tyrosine kinase analysis assay
Protein kinases are analyzed using labeled donor substrates, and the accumulation of markers in protein or peptide acceptor substrates is readily detected when
Experiments for direct analysis of kinases in gels
Protein kinases are analyzed using labeled donor substrates, and the accumulation of markers in protein or peptide acceptor substrates is readily detected when
Fusion Protein Enzymolysis Assay
The use of gene fusion expression systems to express exogenous genes in E. coli has become increasingly popular. The reason for this is largely attributed to th
Chemical degradation of fusion proteins
The use of gene fusion expression systems to express exogenous genes in E. coli has become increasingly popular. The reason for this is largely attributed to th
Solubilization and replication experiments of inclusion body proteins
Although inclusion bodies are mixtures of unnatural forms of expressed proteins, the peptide chain itself is intact, or the primary structure is correct. One ad
Determination of protein concentration by Lowry's assay
The Lowry method is a standardized, rapid method for protein quantification and has been widely used to remove interfering substances by protein precipitation p
Experimental determination of protein concentration by spotted filter membrane binding method
The spotted filter binding method is a rapid method for the initial determination of protein content. This method is particularly useful for screening useful fr
Discontinuous nondenaturing coagulation limb electrophoresis experiments
Non-denaturing gel electrophoresis, also known as natural gel electrophoresis. The separation of a protein in a gel depends on its charge and molecular size. Wh
Capillary electrophoretic analysis of proteins
The main features of capillary electrophoresis are high column efficiency (N＞105-106), short analysis time, low consumption of sample volume and reagents, mult
Batch Chromatography for Protein Concentration
Batch chromatography is simple and fast, suitable for situations where protein resolution is not required, and an effective means of concentrating proteins. Sou
Preparation of affinity columns
This section will focus on the solid-phase technique of ligand solidification on agarose gels activated with cyanogen bromide. The vast majority of ligands, suc
Immunopurification experiments
Antibody affinity purification is one of the most effective methods in protein purification. This step alone often results in 1000 to 10,000-fold purification.
Experiments on protein purification by lectin affinity chromatography
Lectins are proteins that reversibly bind carbohydrates. Because most lectins have at least two carbohydrate binding sites per molecule, they can precipitate gl
Dye-matched base chromatography for protein purification experiments
Dye-ligand chromatography is not really affinity chromatography because they are not natural ligands for the proteins to which they bind. However dye columns bi
Protein expression experiments in mammalian cells
Content from Compact Molecular Biology Laboratory Guide (5th Edition)
Interaction cloning experiments
Interacting cloning (also known as expression cloning) is used to identify and clone proteins encoding proteins that interact with a target or 'bait' protein. T
Experiments on the purification of proteins by ion exchange chromatography
Ion exchange chromatography is the most widely used of the chromatographic means for purifying proteins. It has a high resolution for proteins, is easy to perfo
Experiments on the purification of proteins by filtration chromatography using filter gel
Gel filtration chromatography is an important protein purification technique, also known as size exclusion, gel exclusion, molecular sieve, or gel filtration ch
Dissolution and reduction of samples for two-dimensional polyacrylamide gel electrophoresis
Source : Proteomics: From Sequence to Function.
Phosphorylation Site Analysis Experiment - Determination of Phosphorylated Amino Acid Types
Source : Proteomics: From Sequence to Function.
Lysis experiments of cultured animal cells, yeast and bacteria for immunoblotting
Source: Laboratory Guide to Proteins and Proteomics
Experiments on lysis of cultured cells by depressurization of nitrogen compartments
Source: Laboratory Guide to Proteins and Proteomics
Experiments on bulk extraction of recombinant proteins from bacteria
Source: Laboratory Guide to Proteins and Proteomics
Experiments on lysis of recombinant proteins of Escherichia coli from inclusion bodies
Source: Laboratory Guide to Proteins and Proteomics
Experiments on the preparation of yeast extracts
Source:Laboratory Guide to Proteins and Proteomics
Determination of protein concentration by BCA method
Determination of protein concentration in samples
Total protein extraction from adipose tissue and cells
Extraction of total protein from adipose tissue and cells
Biopharmaceutical Glycosylation Site Assay
N-glycosylation site studies are one of the most important prerequisites for understanding the function of sugar chains
Fluorescent staining of frozen sections for amyloid (AMYLOID) thioflavine T
Fluorescent staining of protein fibers
RIP technology
Studying RNA-protein binding in cells
Far Western analysis of protein interaction experiments
When analyzing protein interactions using Far Western blot, the target protein is immobilized on a solid support membrane and then probed with a non-antibody pr
Affinity purification assay of GST fusion proteins
This experiment describes the application of proteins fused to glutathione-S-transferase (GST fusion proteins) to affinity purify other proteins, also known as
Recombinant MVA in vivo immunostaining assay
In vivo immunostaining can be used for the detection of modified poxvirus Ankara (MVA) and also for the identification of proteins expressed on the cell surface
Protein kinase C isoform analysis experiment
Protein kinases are analyzed using labeled donor substrates, and the accumulation of markers in protein or peptide acceptor substrates is readily detected when
Dialysis for the removal of sodium N-dodecyl sarcosinate
This experiment describes how much time it takes to remove sodium N- dodecyl sarcosinate by dialysis. This experiment is from the Laboratory Guide for Protein P
Measurement of extinction coefficients (Scopes method)
Scopes published a simple method for determining the extinction coefficient (E) of proteins (Scopes, 1974). This method is based on the fact that although most
UV spectra of pure proteins A280 nm/A260 nm Experimental
A relatively simple and effective measure of protein purity is obtained by determining the ultraviolet (UV) spectra of purified proteins. Even if sodium dodecyl
Quantitative SDS gel staining and scanning
Although most proteins bind approximately the same amount of dye per milligram, there is still a 10% or greater variation in the ability to bind dye between pro
Dissolution, refolding and ion exchange chromatography experiments on inclusion body precipitates (σ32)
This experiment describes the solubilization, refolding and ion exchange chromatography of inclusion body precipitation (σ32). This experiment was derived
Carrier amphoteric electrolyte pH gradient isoelectric focusing experiment
Isoelectrofocusing, abbreviated as IEF or EF, was established in the 1960s as a means of separating and analyzing proteins, and has evolved rapidly over the pas
Determination of sucrase activity at various levels and calculation of purification rate experiments
The rate of sucrose hydrolysis was determined by measuring the amount of reducing sugars (glucose and fructose) produced, and in this experiment, the unit of su
DEAE cellulose column chromatography purification of enzyme protein experiment
The aim of this experiment was to further purify sucrase protein by column chromatography, using DEAE cellulose resin as an ion-exchange agent for hierarchical
Formaldehyde denaturing gel electrophoresis to identify RNA experiment
The purpose of this experiment is to learn how to identify RNA by formaldehyde denaturing gel electrophoresis. Agarose gel electrophoresis is often used for the
Extraction of total plant RNA
Grind the plant material in liquid nitrogen, make some cells broken, further make the plant cells lysed in lysis solution, precipitate protein with sodium aceta
Tm value measurement of DNA
When the dilute salt solution of DNA is heated to 80~100 ℃, the double helix structure will be disintegrated, the two strands will be separated, and a random c
Isolation and purification of animal genomic DNA
The purpose of this experiment is to master the basic principles and methods of mass preparation of animal genomic DNA by salt solubilization method, according
Separation of lactate dehydrogenase isozymes by agarose gel electrophoresis assay
Lactate dehydrogenase is present in all glycolytic cells, and catalyzes the dehydrogenation of lactate to form pyruvate or the reduction of pyruvate to lactate
Expression and purification experiments of LacZ and trpE fusion proteins
Fusion proteins are generally used as antigens to stimulate antibody production and in many cases for biochemical analysis. Two widely used E. coli expression s
Heparanase Lowry colorimetric assay
The Lowry colorimetric method is one of the most sensitive methods for protein determination. The color development principle of this method is the same as that
Determination of substrate binding spectra to hepatic cytochrome P-450
The hepatic cytochrome P450 system plays an important role in the metabolism of drugs by converting them from hydrophobic to more easily excreted hydrophilic fo
Extraction and activity of ATPase from cardiac myofibrillar fibers
Cardiac myofibrillar ATPase is Ca2+, Mg2+ ATPase, which breaks down ATP into ADP and Pi and releases energy to provide myofibrillar contraction needs. Myofibril
TNF-α Biological Activity Assay
One of the biological activities of TNF is its ability to directly kill tumor cells.TNF binds to the corresponding receptor and then moves into the cell, where
Reversible staining of proteins on PVDF membranes
Reversible staining of proteins on PVDF membranes can be used for: confirming the transfer of proteins to PVDF membranes during Western hybridization.
Purification of PPO crude enzyme solution
The purification of PPO can be applied to (1) the study of PPO nutritional mechanisms and (2) PPO physiological mechanisms and regulation.
Identification of IgG purity by polyacrylamide gel electrophoresis
Since the concentration of the polyacrylamide gel can be prepared on demand, two electrophoresis systems, the "Continuous System" and the "Discontinuous System,
Experimental determination of protein concentration by bicinchoninic acid (BCA) assay
The bicinchoninic acid (BCA) assay is a new and improved Lowery assay that has recently been developed, with a simple reaction and virtually no influence of int
Dialyzed to obtain soluble refolding σ32 Monomer experiments
This experiment describes what dialysis conditions lead to the highest yield of soluble refolded σ32 monomer. This experiment is from the guide to protein
Elution from PEI precipitation σ32 and RNA polymerase assay
This experiment describes the amount of salt needed to elute σ32 and RNA polymerase from PEI precipitation. This experiment is from the Protein Purificati
Preparation of DNAase I Footprint Probes
DNA enzyme I footprinting analysis is an effective method for detecting and characterizing the sequence-specific binding ability of transcription factors to DNA
Determination of NADPH-cytochrome C (P-450) activity
NADPH-Cytochrome C (P-450) is a heme protein that is a terminal oxidase due to the fact that when it is in its reduced form, it combines with carbon monoxide to
Experiment to determine the content of cell membrane proteins
Membrane proteins are protein molecules present in the cytoplasmic membrane of cells and play important roles in intracellular molecular transport, energy metab
Determination of total protein content based on protein elemental composition
Determination of total protein content based on the elemental composition of proteins is a method of direct analysis based on the characteristics of the element
Determination of total protein content based on protein light absorption properties
Determination of total protein content based on protein light absorption properties is based on the light absorption properties of different chemical bonds of p
Experiments for precise molecular weight determination of proteins
The exact molecular weight of a protein is the sum of all the atomic weights contained in the molecule. Currently, there is one main method used to determine th
Experiments for the determination of the isoelectric point of proteins
The isoelectric point of a protein is one of the important properties of proteins. Determination of isoelectric point of proteins is important for understanding
GST pull-down analysis
Protein-protein interactions play an important role in a variety of life processes such as biocatalysis, transporter, signaling, immunity, cellular regulation,
X-ray crystallography
The ability of protein molecules to crystallize has been reported for more than 170 years, and the discovery of X-rays by Rntgen WC in 1895 and X-ray diffractio
Determination of key enzymes and important products in the anaerobic oxidation pathway
Glucose undergoes a 10-step metabolic reaction of glycolysis to produce keto acid, which is then further produced as lactate or decarboxylated into acetyl coenz
Pentose phosphate pathway
The pentose phosphate pathway is an oxidative catabolism of glucose that produces NADPH and synthesizes ribose phosphate, which provides the raw material for nu
Lipid metabolite analysis
Lipid metabolism is an important source of energy supply for the body, and an imbalance between lipid synthesis and catabolism triggers lipid metabolism disorde
Analysis of amino acid metabolites
Amino acids are mainly used in the body to synthesize proteins, peptides and other nitrogenous substances that are unique to the body; on the other hand, they c
Electrophoretic separation of proteins
Proteins are amphoteric electrolytes and exhibit charged properties in solutions above or below the isoelectric point. These charged proteins migrate by swimmin
Yeast two-hybrid technology
Yeast two-hybrid (yeast two-hybrid) is an experimental system for studying the effects of interactions between trans-acting factors on the transcriptional regul
Protein Oligomerization Assay
Oligomerization refers to the formation of dimers, trimers, tetramers or long chain molecules (oligomers) between monomeric compounds. In a biological context,
Acid phosphatase activity assay
Acid phosphatase (ACP) is widely found in various cells, and is often abnormally expressed in some cancer cells. Sodium p-nitrophenol phosphate is used as the substrate of ACP, which can be hydrolyzed under alkaline conditions to produce yellow p-nitrophenol, and ACP can be quantitatively ...
Dual luciferase reporter assay
The target gene transcriptional regulator original is constructed onto the firefly luciferase reporter gene plasmid, and co-transfected with the sea kidney luci
Eukaryotic expression of proteins
A protein expression system is a system consisting of a host, an exogenous gene, a vector, and auxiliary components. The vector contains an exogenous gene fragm
Protein expression system construction
E. coli protein expression is a commonly used protein expression system. E. coli is a common bacterium, which has been widely used in the field of bioengineerin
Activation and monitoring of inflammatory vesicles
Nucleotide-binding oligomerization domain-like receptor family, pyrin domain-containing 3 (NLRP3) inflammatory vesicles have been well studied in recent years.
Isolation and culture of osteoclasts
As highly differentiated multinucleated giant cells, osteoclasts are mainly derived from monocyte/macrophage hematopoietic stem cell line, which is a kind of ce
Angiogenesis experiment
In the process of tumor development, tumor tissues are often in a state of hypoxia and low nutrition. Under such conditions, due to the activation of angiogenic
Protein purification experiments
His-Tag is the most common tag used in the purification of recombinant protein expression and can be used for initial purification of most proteins (soluble or
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