Magnetic Beads and Applications
Affinity Magnetic Beads
  • Amine
  • Aldehyde
  • C arboxy l
  • C DI
  • D VS
  • D ADPA
  • E poxy
  • H ydrazide
  • H ydroxyl
  • I odoacetyl
  • N HS
  • S ulfhydry
  • T osyl
  • T hiol
  • S ilica
  • IDA
Reversed-Phase  Beads
  • C 4
  • C 8
  • C 18
  • Cyanopropyl
  • Phenyl
  • di Phenyl
Ion Exchange Beads
  • D EAE (WAX)
  • P SA (WAX)
  • S AX
  • W CX
  • S CX
  • Hydroxyapatite
Antibody Purification & IP
  • P rotein A
  • P rotein G
  • P rotein A /G
  • P rotein L
  • Q uick IgG Pure Beads
  • A ntigen Peptide
  • Q uick IgM Pure Beads
  • A nti-IgG Beads
  • Q uick Ig A Pure Bead
  • T hiophillic Beads
Antibody Immobilization
  • P rotein A
  • P rotein G
  • P rotein A /G
  • P rotein L
  • E poxy
  • Aldehyde
  • H ydrazide
  • C arboxyl
  • I odoacetyl
  • T hiol
Recombinant Protein
  • N i+ Charged Beads
  • C o + Charged Beads
  • M altose
  • Calmodulin Beads
Peptide Immobilization
  • E poxy
  • Aldehyde
  • C arboxyl
  • Amine
  • I odoacetyl
  • T hiol
DNA/RNA Purification
  • B lood
  • S aliva
  • S tool
  • F ood
  • S oil
  • P CR Products
  • m RNA
  • Heteroduplex DNA
    (Mutation Isolation)
DNA/RNA Immobilization
  • G enomic DNA/RNA
  • m RNA
Endotoxin Removal
Small Molecule
  • E poxy
  • Aldehyde-terminated
  • H ydrazide -terminate d
  • D VS-activated
Abundant Protein Removal
  • H SA Removal
  • I gG Removal
  • H SA/IgG Removal
  • A bundant-10 Protein
  • A bundant- 2 2 Protein
Cell Isolation
  • V irus
  • B acteria
  • C ell
  • P arasites
EDTA-Magnetic Beads
  • N TP /Oligonucleotides
  • E nzyme
  • A ntibody
  • P eptide
  • B oronic Acid
  • Disulfide Reductants
I DA-Magnetic Beads
Recombinant Protein
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Recombinant Protein & cDNA
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Biomagnetic Applications For Bioseparation
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Recombinant Proteins
In the proteomics era, there are increasing demands and expectations on recombinant protein production for functional and
structural studies. Many laboratories face challenges in recombinant protein production, particularly proteins that are poorly
characterized. After identification of the protein/gene sequence of interest, the first step is to obtain the cDNA for cloning.
There are two ways of doing that: one is to obtain the DNA or mRNA from the species where the protein originates; the
other is to make it by chemical synthesis, which is more convenient particularly if the native cDNA is not readily available. If
DNA/mRNA is obtainable from the original species, PCR is the method of choice to amplify the target cDNA, and to add
cloning sites for cloning into an appropriate vector. Another advantage of the synthetic method is that you may design the
nucleotide sequence based on the protein sequence. This is particularly useful if subsequent expression of a eukaryotic
protein is to be carried out in prokaryotic systems like E. Coli. There are some codons that are more frequently used in
eukaryotic organisms but are rarely used in lower organisms. For example, if there are some codons in a human gene that
are rarely seen in E. Coli, expression using the original cDNA sequence would likely result in lower yield. In such cases, we
may design cDNA sequences based on usage of E Coli codons.

   A very important consideration before cloning is the choice of an expression system and an expression vector for that particular
organism. There are two major expression systems, either prokaryotic (E. Coli. etc.) or eukaryotic (yeast, insect cells, plant cells, or
mammalian cells). The choice of the expression system will decide which vector you will use to clone your cDNA because different
organisms use different promoters to drive expression of the recombinant protein. Although vectors with multiple promoters that work in
different organisms have been developed, they usually do not work as efficiently as vectors with a single promoter. If relevant
information is already known about the target protein from published literature or can be deduced from similar proteins that have been
successfully produced, it’s easy to make a decision on the expression system and the vector. However, if little is known concerning
the target protein, multiple systems may be tried in parallel if time is a major concern, or in most cases, the prokaryotic (E Coli)
system would be tried first. If expression level is low or the protein is not soluble in E. Coli, then try the eukaryotic system (typically
insect cells or mammalian cells). An alternative is to attempt refolding of insoluble proteins from E. Coli, but it is usually more time
consuming and often prove to be a failure. There are some general considerations in deciding which system or vector to use.
 The prokaryotic system is easy to manipulate, bacteria grow more rapidly, expression is easily induced by IPTG, and recombinant
protein purification is relatively simple. But E. Coli usually is unable to handle large or more complex proteins, solubility is also an
issue, particularly with membrane proteins, and some post-translational modifications are absent in E. Coli, which make it
inappropriate for functional or enzymatic studies. In these cases, the eukaryotic system is preferred. There are eukaryotic vectors that
contain secretory signals which secrete the recombinant protein into the culture medium. You may keep growing the cells
continuously and collecting the medium from which to purify the protein. But eukaryotic cells grow relatively slowly. Usually a strong
promoter is used in an expression vector to produce more mRNA. Commonly used promoters in E. Coli vectors include T7, lambda
P1, and araB. The popular pET vectors are based on T7 promoters. However, leaky expression of T7 RNA polymerase can cause
instability of plasmid. Repression of the leaky expression is achieved by a lac operator and T7 lysozyme (pLysS). E. Coli strains
optimized for recombinant protein production are available from commercial suppliers.

 Baculovirus mediated recombinant protein production in insect cells has become more and more popular for recombinant protein
production especially in structural and functional studies. Insect cells offer post-translational modifications similar to mammalian cells.
This system uses a helper-independent virus which can be amplified to extremely high titers. The insect cells which are adherent can
easily be adapted to suspension culture. A number of baculovirus vectors are available for recombinant protein expression. In most
cases, the target gene is first cloned into a transfer vector that contains sequences flanking the polyhedron gene of the baculovirus
genome. After co-transfection into the insect cells together with the baculovirus genome, the target gene is transferred into the virus
genome via homologous recombination. The strong late polyhedron promoter drives expression of the target gene. Another more
efficient method to generate recombinant virus is by site-specific transposition of the target gene in a plasmid into a bacterial artificial
chromosome (bacmid) which contains the infection and replication elements of the baculovirus, which may reach high titer after
amplification in insect cells. This is commercially available from Invitrogen (Bac-to-Bac). There are a number of other approaches to
make recombinant baculoviruses. The most frequently used insect cell lines include Sf9 and Sf21, both derived from the pupal ovarian
tissue of S. frugiperda. Highh Five is another commonly used insect cell line derived from Trichoplusia ni. There are a number of other
insect cell lines claimed to offer advantages over Sf9 or Sf21, but in most cases, Sf9 and Sf21 are typically used in routine
recombinant protein production. It is noted that the glycosylation profile in insect cells is not identical to that in mammalian cells.

Yeast is an important system for recombinant protein production in research, industrial and medical applications. Yeast can be grown
rapidly in simple medium to very high density, thus making it ideal for large scale culture. And it is genetically easy to manipulate
similar to E. Coli. As a single cell eukaryote, particularly Pichia pastoris, provides a powerful tool for production of secretory
recombinant proteins.

Recombinant protein production in Mammalian cell  have obvious advantages over the other systems, including appropriate post-
translational modification and correct folding. There are two approaches for mammalian expression: stable cell line and transient
expression. The establishment of stable mammalian cell lines is often labor and time consuming, whereas, the transient system
provides an option for production of recombinant proteins in mammalian cells in a shorter time. CMV is the most commonly used
promoter in transient expression vectors.

Cell-free expression systems offer an alternative for quick production of recombinant proteins, but its application has been hindered by
low expression levels. Recent improvements have resulted in availability of several commercial kits, but they are quite expensive.

Plant expression systems offer eukaryotic post-translational modifications, are easy to scale up, and free of bacterial or viral
contaminations. There are two main strategies for plant expression of recombinant proteins: transgenic and transient. The transgenic
approach inserts the target gene into the nuclear or chloroplast genome, whereas, the transient approach uses a plant virus to
introduce the target gene into the plant cells. The transgenic strategy takes a longer development time in contrast to the faster and
more cost-efficient transient expression system.

In most cases, a tag is added to the recombinant protein for purification or other purposes. Commonly used tags for recombinant
protein purification purpose include GST, polyhistidine (6xHis), MBP, Inteim CBD, etc. Purification kits are commercially available from
suppliers. These tags can be placed either at the N- or C-terminus of the target recombinant protein. Some tags may also help to
increase the expression level or solubility of the fusion recombinant protein. Each tag seems to offer some advantages, and thus it is
difficult to decide which to choose. Sometimes more than one tag is used in combination to obtain the best result. If removal of the tag
is desired, a protease cleavage site may be incorporated at the N-terminus of the recombinant fusion partner. Commonly used
cleavage sites are Enterokinase, Factor Xa, Thrombin, TEV protease, and PreScision protease. There may be problems with the use
of proteases, including incomplete cleavage, resulting in reduction of the protein, or non-specific cleavage resulting in heterogeneity in
the protein.
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Monoavidin Magnetic Beads
Features and Advantages:
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His-tagged Recombinant Protein
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Quick DNA & RNA Purification
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C4, C8, C18 Magnetic Beads
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Quick Peptide Conjugation &
Antibody Purifiction
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Protein A, G, A/G, L Magnetic Beads
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