ImmoMix™ is a complete ready-to-use, red-colored, high yield; heat-activated, 2x reaction-mix which simply requires the user to add water, template and primers, and then pre-heat to 95°C for 10 minutes to carry out successful PCR assays. Once completed, the reaction mix can be loaded directly on to a gel.
ImmoMix™ Red is a complete ready-to-use heat-activated 2x reaction-mix, which simply requires the user to add only water, template and primers, and then pre-heat to 95°C for 10 minutes to successfully carry out PCR assays. The 10 minute activation step eliminates the presence of non-specifics such as primer-dimers and mis-primed products, since the enzyme is inactive at initial low temperatures.
ImmoMix Red combines all of the features and advantages of ImmoMix and contains an additional inert red dye. This non-toxic, non-hazardous red dye allows users to load samples directly onto a gel, without the need to add loading buffer since the mix is of sufficiently high density to sink to the bottom of the gel.
Adequate mixing is also ensured when reactions are set up. The red dye migrates like a 350 bp fragment on a 2% agarose TAE gel (or 600 bp on a 1% agarose).
ImmoMix Red is based on IMMOLASE™ DNA Polymerase, which leaves an ´A´ overhang and can be used for a wide variety of templates. An additional 50 mM MgCl2 solution is included should any fine adjustments be required.
ImmoMix Red reduce the time needed to set up reactions, thereby reducing the risk of contamination. Greater reproducibility is ensured, by reducing the number of pipetting steps that can lead to pipetting errors.
10 x 1.25 mL
50 mM MgCl2 Solution
All components should be stored at -20°C upon receipt for optimum stability. Repeated freeze/thaw cycles should be avoided.
When stored under the recommended conditions and handled correctly, full activity of the reagents is retained until the expiry date indicated on the outer box label.
Components may also be stored at +4°C and will remain stable for up to 2 weeks, however storage at -20°C is recommended.
On Dry Ice or Blue Ice.
Bioline's mixes contain magnesium chloride at the following concentrations (please see table below). An additional tube of 50 mM MgCl2 solution is supplied with each mix to facilitate further optimization if required.
|Bioline Mix||Final Magnesium Concentration|
|ACCUZYME Mix||2.0 mM.|
|BioMix / BioMix Red||2.5 mM.|
|ImmoMix / ImmoMix Red||3.0 mM.|
|BIO-X-ACT Short Mix||2.0 mM.|
IMMOLASE DNA Polymerase requires a heat-activation step of 10 minutes at 95°C.
As well as most standard applications, IMMOLASE DNA Polymerase is ideally suited to the following:
- High-throughput applications
- Multiplex PCR
- TA Cloning
IMMOLASE DNA Polymerase has been manufactured under 13485 Quality Management System and is suitable for further manufacturing use as an IVD component.
The features of IMMOLASE DNA Polymerase are as follows:
Optimal Extension Temperature: 72°C
|No or low PCR yield||Enzyme concentration too low – increase the amount of enzyme in 0.5 U increments.|
|Primers degraded – check quality and age of the primers.|
|Magnesium concentration too low – increase concentration in 0.25 mM increments with a starting concentration of 1.75 mM.|
|Primer concentration not optimized. Titrate primer concentration (0.3-1 µM); ensuring that both primers have the same concentration.|
|Template concentration too low – Increase concentration of template.|
|Perform a positive control to ensure that the enzyme, dNTPs and buffers are not degraded and/or contaminated.|
|Multiple Bands||Primer annealing temperature too low. Increase annealing temperature. Primer annealing should be at least 5°C below the calculated Tm of primers.|
|Prepare master mixes on ice or use a heat-activated polymerase.|
|For problems with low specificity. Try adding 3% DMSO (not supplied) to improve specificity.|
|Smearing or artifacts||Template concentration too high. Prepare serial dilutions of template.|
|Too many cycles. Reduce the cycle number by 3-5 to remove non-specific bands.|
|Enzyme concentration too high - decrease the amount of enzyme in 0.5 U increments.|
|Extension time too long. Reduce extension time in 0.5-1 minute increments.|