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Isolation and analysis of c-myc-tagged proteins
Background
The c-myc peptide (EQKLISEEDL) originates from the human c-myc proto-oncogene, which is overexpressed in a wide range of human cancers. It has become a popular fusion tag for mammalian recombinant protein expression.

A high-quality monoclonal Anti-c-myc antibodies is offered by Miltenyi Biotec either coupled to µMACS™ MicroBeads for specific isolation of c-myc-tagged proteins or conjugated to biotin, FITC, or horseradish peroxidase (HRP) for analysis of the recombinant c-myc-tagged protein.
Isolation and detection of c-myc-tagged proteins
Isolation of c-myc-tagged proteins

The µMACS™ and MultiMACS™ c-myc Isolation Kits take advantage of the well-established MACS® Technology; thus, allows fast and specific isolation of pure c-myc-tagged proteins (fig. 1). The advantages of MACS Technology for protein isolation are:

Specificity – high-quality monoclonal antibodies coupled to MACS MicroBeads assure specific isolation of c-myc-tagged proteins with low background.
Sensitivity – fast reaction kinetics of tiny (50 nm), non-sedimenting MACS MicroBeads allow sensitive isolation even when working with rare proteins.
Speed – less than 2 hours to high-purity c-myc-tagged proteins!

The µMACS c-myc Isolation Kit was developed for manual, low-throughput applications with the µMACS Separator.
The procedure can easily be upscaled with the MultiMACS c-myc Isolation Kits to a semi- or fully-automated, high-throughput processing of up to 96 samples in parallel by utilizing the MultiMACS 96 Separator.


Detection of c-myc-tagged proteins

The monoclonal Anti-c-myc antibody is available with various conjugates:
The FITC-conjugated Anti-c-myc antibody enables an immunofluorescence analysis, e.g. by fluorescence microscopy (fig. 2). Directly coupled to horseradish peroxidase (HRP), the Anti-c-myc-HRP antibody simplifies immunological methods, as incubation with secondary antibodies is not necessary. The Anti-c-myc-HRP antibody was generated for specific and sensitive detection of c-myc-tagged proteins by Western Blot (fig. 3) or ELISA analysis. The biotinylated Anti-c-myc antibody can be used for various detection methods in combination with appropriate streptavidin conjugates.

Target sequence: EQKLISEEDL
Tag origin: Human c-myc protooncogene
Isotype: mouse IgG1
Columns
For µMACS™ Isolation Kit: µ Column, upscale with M Column

For MultiMACS™ Isolation Kits: Multi-8 or Multi-96 Columns
Further information
Protein research brochure.pdf
[PDF; 850,1 KB]
 
Figure 1
Specific isolation of recombinant fusion protein with μMACS™ Anti-c-myc MicroBeads. Cells were transfected with a vector encoding c-myc-BDCA-2 and in vivo labeled with 35S-methionine. The purification of recombinant c-myc-BDCA-2 protein was performed using Anti-c-myc MicroBeads and, as a negative control, Anti-HA MicroBeads. The figure shows an autoradiogram after SDS-PAGE.
Figure 2
Immunofluorescence analysis of c-myc-BDCA-2 expressing CHO cells. CHO cells were transiently transfected with a vector encoding c-myc-BDCA-2 and analysed by fluorescence microscopy after 24 hours of expression. A Detection with Anti-c-myc-FITC and CD303 (BDCA-2)-Biotin/Anti-Biotin-Alexa 546. B Detection with Anti-c-myc-Biotin/Anti-Biotin-Alexa 546 and CD303 (BDCA-2)-FITC; red: Alexa 546-labeling, green: FITC-labeling, yellow: FITC-Alexa 546 double-labeling.
A
B
Figure 3
Sensitive detection of recombinant fusion protein with Anti-c-myc-HRP antibody. 293 HEK cells were transfected with a vector encoding c-myc-BDCA-2. 5×106 cells were lysed and recombinant protein c-myc-BDCA-2 was purified using the μMACS™ Anti-c-myc Isolation Kit. 1/100, 1/500, and 1/1000 of the eluate was separated by SDS-PAGE, blotted on a PVDF membrane and detected using Anti-c-myc-HRP (1:10,000, 1 h, room temperature) and ECL reagents (GE Healthcare).
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Details
Products
μMACS Anti-c-myc Starting Kit
- for 40 isolations
Components
- 1 μMACS c-myc Isolation Kit
- 1 μMACS Separation Unit
- 1 MACS MultiStand
- 2×20 μ Columns
130-091-284
Qty.:
 

μMACS c-myc Isolation Kit
Tag specification
- for 40 isolations
Components
- 2 mL μMACS Anti-c-myc MicroBeads
- 2×50 mL Lysis Buffer
- 50 mL Wash Buffer 1
- 5 mL Wash Buffer 2
- 5 mL Elution Buffer
Download datasheet
130-091-123
Qty.:
 

MultiMACS c-myc Isolation Kit (12×8)
- for 96 isolations
Components
- 3x2 mL µMACS Anti-c-myc MicroBeads
- 1x50 mL Equilibration Buffer
- 1x Multi-8 Column Box (12x8)
Download datasheet
130-094-250
Qty.:
 

MultiMACS c-myc Isolation Kit (4×96)
- for 384 isolations
Components
- 5×4.6 mL µMACS Anti-c-myc MicroBeads
- 1×50 mL Equilibration Buffer
- 1x Multi-96 Column Box (4x96)
Download datasheet
130-094-251
Qty.:
 

Anti-c-myc-HRP
Specification
- 100 μL
Download datasheet
130-092-113
Qty.:
 

Anti-c-myc-FITC
Product Specifications
- 1 mL
- for up to 200 tests (intracellular immunofluorescent staining)
Download datasheet
130-092-472
Qty.:
 

Anti-c-myc-Biotin
Product Specifications
- 1 mL
- for up to 200 tests (intracellular immunofluorescent staining)
Download datasheet
130-092-471
Qty.:
 

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Protein-Protein- / Protein-DNA-Interaction
Isolation and analysis of GST-tagged proteins
μMACS™ and MultiMACS™ Streptavidin Kits
MACSelect™ - Transfected Cell Selection Kits
References
1. Indran et al. (2010) J. Virol. 84: 3162-3177.
2. Staubach et al. (2009) Proteomics 9(10): 2820-2835.
3. Robert et al. (2006) J. Biol. Chem. 281(52): 40135-40143.
4. Spoelgen et al. (2006) J. Neurosci. 2: 418-428.
5. Mach et al. (2005) J. Virol. 79: 2160-2170.
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