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ImmoMix™

Ordering

Cat. No.
Size
BIO-25020
500 x 50µl Reactions

*link will take you to our exclusive distribution partner site

Description

ImmoMix™ is a complete ready-to-use 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.

Product Highlights

  • Convenient - pre-mixed, pre-optimized 2x solution
  • Ready to use format - reduces risk of contamination
  • Fast set up - decreased time compared to traditional methods
  • Reproducible results - time after time

Product Description

ImmoMix™ is a complete ready-to-use 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 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 is based on IMMOLASE™ DNA Polymerase, which leaves an ´A´ overhang, and has been optimized for a wide variety of templates. Additional MgCl2 solution is included should any fine adjustments be required.

ImmoMix reduces 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.

Applications

  • Ultra-high specificity for multiplex reactions
  • Products suitable for TA cloning
Main

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Specification

Components

Reagent

500 Reactions

ImmoMix

10 x 1.25 mL

50 mM MgCl2 Solution

1.2 mL

Volume

  • BIO-25020: 500 x 50 µL Reactions: 10 x 1.25 mL

Concentration

  • 2x

Storage & Stability

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 on the outer box label.

Components may also be stored at +4°C if required, although storage at -20°C is recommended. When stored at +4°C, reagents will remain stable for a period of two weeks from date of receipt.

Shipping conditions

On Dry Ice or Blue Ice.



Resources

Documents

PCR Enzyme Guide - A New Generation of End-Point PCR
ImmoMix - MSDS
IMMOLASE DNA Polymerase - Fact Sheet
ImmoMix - Product Manual

Latest Certification of Analysis (COAs)

BIO-25020_PM354-B124150

FAQs

What is the final working concentration (at 1x) of MgCl2 in each mix?

Meridian'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.



 Meridian 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.
MangoMix  2.5 mM.
MyTaq  3.0 mM.
RANGER  1.5 mM.



What is the heat activation time required for this polymerase?

IMMOLASE DNA Polymerase requires a heat-activation step of 10 minutes at 95°C.



Which specific applications is IMMOLASE DNA Polymerase suited to?

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.



What are the features of IMMOLASE DNA Polymerase?

The features of IMMOLASE DNA Polymerase are as follows:

  • Heat Activation: 10 minutes at 95°C
  • Speed: 15-30 s/kb (template dependant)
  • Length amplified: Around 5 kb (genomic DNA)

Optimal Extension Temperature: 72°C



Which polymerase do I need for my specific application?

At Meridian we pride ourselves in supplying high-quality polymerases to suit your requirements. To aid your selection of the most suited enzyme for your specific applications, please see our enzyme selection tool.

What are the storage conditions and stabilities of Meridian polymerases?

All our polymerases are guaranteed for a period of 12 months from the date of purchase. These should be stored at -20°C during this time for optimal retention of activity.

Please Note: We do not recommend the storage of our polymerases at -80 °C as ice crystals could form on the active site, which may affect or destroy the activity of the enzyme.

I am having problems optimizing my PCR, what would you recommend?

PCR can be a challenging technique, with various parameters to optimize to achieve the best results. If you are having problems, these could be easily resolved by addressing a few issues. Please see our PCR troubleshooting guide for suggestions and help with your specific problems.

Observation Recommended Solution(s)
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.


What do the terms yield, fidelity, processivity and specificity actually mean?

These terms refer to parameters to be considered when performing PCR and are important features in selecting the correct enzyme for your needs. Understanding what they mean is therefore crucial: Yield: The amount of DNA produced in a PCR reaction. Fidelity: The accuracy of the enzyme at incorporating the correct dNTP to the elongating DNA strand. Processivity: The length of time a polymerase is associated with the template and therefore the size of fragment which can be amplified. Specificity: A measure of the unwanted by-products generated in a reaction.

What are the benefits of using a hot-start polymerase?

During PCR setup at room temperature, standard polymerases will have some activity. When using a hot-start polymerase, this enzyme will have no activity at room temperature, thus reducing the risk of unspecific products in the reaction due to these mis-primed oligonucleotides. This feature makes this type of polymerase ideally suited to high-throughput applications, where the reactions may be left sitting at room temperature for prolonged periods of time.

What is a proofreading polymerase?

In nature, the ability of a DNA polymerase to correct misincorporations of nucleotides in the DNA strand being elongated is often crucial to the survival of the host organism. This ability is termed "proofreading activity" and occurs in the 3' to 5' direction. This activity also leads to the polymerase removing unpaired nucleotides overhanging at the 3'end (A-overhangs), creating blunt ends.

How is one unit of activity of a polymerase defined?

One unit is defined as the amount of enzyme that incorporates 10 nmoles of dNTPs into acid-insoluble form in 30 minutes at 72°C.

What are the advantages of using a 2x mix rather than setting up a PCR reaction from scratch?

All Meridian polymerases are available in the convenient format of a 2x master mix, containing all the reagents and additives necessary to perform successful PCR, with the exception of template, primers and water. This formulation not only provides a convenient and hassle-free PCR setup, but also significantly reduces the chances of human error, inaccuracies and contamination by reducing the pipetting steps required.

What is the recommended reaction volume of the polymerase mixes?

We recommend a final reaction volume of 50 µL, this requiring the use of 25 µL of the 2x master mix and to make up to 50 µL using template, primers and PCR grade water.

What is the composition of the polymerase mixes?

Whilst the exact composition of the mixes is proprietary information, all mixes are made up of reaction buffer, magnesium, dNTPs, polymerase and additives, designed to keep the polymerase stable in the mix, as well as provide the optimal conditions for PCR.

What is the concentration of the polymerase mixes?

All Meridian PCR mixes come supplied at a 2x concentration, requiring the user simply to add template, primers and PCR grade water to a final concentration of 1x.

My DNA sample contains PCR inhibitors, which impairs my PCR results. What can I do for optimization?

If possible, we would recommend an additional cleanup of affected samples, for instance, customers had very good experiences with the clean-up of samples containing humic acids with SureClean Plus. Alternatively decreasing the template concentration will help to minimize the amount of inhibitors. The amount of primer can be increased, but do not exceed the suggested limits.

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