YOU NEED TO PASS YOUR DMRQA… THE DATE IS RAPIDLY APPROACHING…WANT A LITTLE HELP?

You’re in charge of the Membrane Filtration Station for microbiology for the DMRQA test this year.

It’s a big responsibility, and demands your fullest attention.

Maybe you’re a new technician, or maybe the person who ,”always does it” isn’t around anymore. What if, you don’t pass, and wouldn’t it be great to have another guide?

This short informative report will help give you some guidance to assure you’re on the road to success when it comes to testing using membrane filtration.

 ONE:

THE 5 “P’S” – LET’S START THERE!

You’ve heard it before:

“Proper preparation prevents poor performance.” Sometimes with colorful language added…but nevertheless it’s a phrase that is worth repeating!

When we talk about being properly prepared in membrane filtration, it begins at the sink. You want to make sure that the filter funnels are nice and clean and free from any debris. They should also be checked periodically for high or low pH walls with a pH indicator solution like bromothymol blue. Make sure all filter funnels, magnetic bases, and manifold are scrupulously clean.

 TWO:

FUNNELS, FLUIDS, AND BBQ BACTERIA!

 After thoroughly cleaning, inspect your polysulfone or glass filter funnel for any deep nicks or scratches. This can harbor bacteria and skew your count or ruin your sterile control.

Sterilize the funnel(s) initially in an autoclave, being sure not to touch the inside walls of the funnel when handling.

TIP: Beware - The funnels come out of the autoclave HOT!

Let them cool, or dispense some sterile dilution water down the interior walls of the funnel prior to filtering the sample so as not to cause any flash thermal exposure (B-B-Q!) to the bacteria, especially if filtering small sample volumes.

Be sure to rinse the interior walls of the funnel after sample filtration with sterile, and properly buffered dilution water. Typically it will be slightly basic.

THREE:

INCUBATORS: “THIS ONE’S TOO HOT, THIS ONE’S TOO COLD, THIS ONE’S JUST RIGHT!”

Different bacteria require specific temperatures for optimum growth. Total Coliform are fine at 35 degrees Celsius, while E. coli – a part of the Coliform Group are thermotolerant and like it hot – at 44.5. degrees C.

When incubating bacteria, because it’s your job to grow them, be sure that the incubator is at the correct temperature not only during the thermometer check times, but use a continuously monitoring device, (plugs into your laptop), so you know for sure that the incubator held temperature during the whole incubation period.

Use a thermometer that has been calibrated against a NIST traceable thermometer.

When laying your membrane filter on the prepared media plated or other media source, place one portion of the filter at an angle onto the media surface – then while applying a very slightly downward pressure simply roll the rest of the filter onto the media surface using your sterile forceps or clips.

FOUR:

TIP: Do at least 5 serial dilutions, more if you have the sample material.

Be sure to incubate your cultures upside down – media side up, to keep the moisture level of the media up.

Following these simple rules will go a very long way in assuring you get great results with your DMRQA test in membrane filtration! Good Luck!

“Membrane Filtration is used the world around – and for very good reason… It works!”

Check out MIDSCI’s pre-poured plates

. 

Improving your Western Blots!

MIDSCI now carries new Expedeon® RunBlue™ Pre-Cast gels!

Check out our options to fit your unit today!

Seven tips to improving your Western Blots:

·         1. Uniformly distributed high background?

o   You may have insufficiently washed.  Increase the number of washes and buffer volume. 

§  Make washing easier with Blot Boxes.

o   Antibody concentrations too high

§  Make sure to optimize your antibody concentrations by running strips at varying concentrations to determine best concentration for your samples.

·         2. Weak or no signal?

o   Your protein may have not transferred well. 

§  Check your transfer buffer make-up and blotting procedure.  Stain gel to visualize proteins remaining in gel. 

·         3. The Ladder Lane is black

o   The antibody may be reacting with your MW (ladder) marker.

§  Add an empty lane between the MW marker and sample lane.

·         4. The band of interest is too low/too high on the blot.

o   Separation is not efficient. 

§  Change the gel percentage: A higher percentage for small protein, a lower percentage for larger proteins. Or try a gradient gel.

·         5. Ripped resolving gel when moving to the cassette for transferring?

o   Try Run Blue™ pre-cast gels from MIDSCI.  They are stronger and resist tearing! No more piecing together torn gels!

·         6. Sample bleeding over to another well or difficulty loading?

o   Try the deeper wells from Expedeon!  Easily hold 35ul of sample without bleeding over to another well!

o   Try gel-loading tips.  Available in flat or round, they make gel loading easier!

·         6. Worried about pre-cast gels expiring or losing resolution before you can use them?

o   Expedeon Run Blue™ gels have a 2 year shelf life with no loss in resolution.  Use them confidently from purchase date to 2 years later!

MIDSCI offers Quality and Affordability with new Saliva Kits!

   Did you know MIDSCI now carries Genefix Saliva kits? 

Besides a more affordable, yet high quality option, how do our Saliva kits compare to others on the market?  Check it out below and please contact us with any questions!

Genefix Saliva Collectors from MIDSCI contain the stabilization buffer in the tube, allowing for rapid sample stabilization that is maintained at room temperature for at least 12 months.  The unique funnel design means no buffer flow back or spillages and the easy to use design allows for use in the field, at home, or clinic based sampling.  The collection tube, certified to 95 kPa, allows for leak-proof mailing.  Saliva DNA Isolation kits are available in both column based and buffer (no column based kits) allowing for both manual and high throughput isolation applications. 

                Want to see the results for yourself?  Contact us for a sample or to purchase a kit at custserv@midsci.com.  For any technical questions, please feel free to e-mail elise.ambrose@midsci.com. 

Choosing a Latex or Nitrile Glove for your needs

Gloves are an essential part of most laboratories.  4 criteria must be met for high quality gloves:

·         Excellent tactile sensitivity

·         Provide barrier protection

·         Remain non-reactive when in contact with patient or sample

·         Provide comfort over extended wear times

MIDSCI™ carries high quality nitrile and latex gloves.  With all of MIDSCI™’s extensive choices, you will not only be able to find a glove that fits your needs, but it will be a glove that you love!

When picking either latex or nitrile gloves, it is important to know exactly which qualities are most important to you.  MIDSCI™ carries 4 main types of latex gloves:

• · Luminance™
• ·  Distinct™
• ·  Velocity™
• ·   Vibrant™
•     Sizes from extra-small to extra-large
•     All of the gloves are manufactured by Aurelia® and undergo air testing for holes to ensure you receive high quality gloves. 

Worried about tearing? Distinct and Velocity are your best latex options while maintaining great elasticity.  The Velocity™ is also Chlorine washed.  The Chlorination ensures the user can get the latex glove on easily without powder. 

Worried about a latex allergy? The Vibrant™ latex gloves are triple washed to significantly reduce the risk of a latex allergy with super low protein content. 

Need excellent grip?  The polymer coating found in the Velocity™, Distinct™, and Luminance™ allows for optimum comfort and increased gripping actions.  Many of the gloves are also textured to ensure maximum grip. 

Are you instead looking for Nitrile gloves?

MIDSCI™ carries Aurelia®’s extensive line of Nitrile gloves. 

Aurelia®’s nitrile gloves offer:

·         Superior comfort and durability

·         Finger texture

·         Air tested to detect pinholes

·         Extensive quality control check to ensure optimum product

Nitrile gloves are offered from the thinnest nitrile available on the market (Amazing™) to a thicker nitrile glove for superior protection (Robust™). 

MIDSCI™ carries 6 main types of nitrile gloves:

·         Amazing

·         Perform

·         Transform

·         Quest

·         Protégé

·         Robust

  

Are you looking for high tactile sensitivity?  We would recommend any of the thin nitrile gloves for this.  Amazing™ gloves are the thinnest, while Perform™ gloves offer strength comfort and sensitivity.

Are you looking for high tactile sensitivity while maintaining durability?  Transform™ gloves are thin yet durable.

Are you looking for a glove similar to latex in comfort, fit and cost, but need nitrile durability and latex-free?  Try the Aurelia® Quest™ glove! Quest™ gloves also maintain great chemical resistance and are finger textured for optimum gripping.  The Protégé™ glove also rivals latex in comfort and ergonomics and comes in a powder blue color.

Need a thicker nitrile glove for improved puncture resistance?  The Robust™ offers a 5 mil thickness and micro-textured finish for improved gripping action. 

MIDSCI™ has a glove to fit your needs.  Contact us today at 1-800-227-9997 for a quote!  For a limited time only, we are offering Buy 15 get 5 free (of the same family)!

PCR plates and strip tubes designed to reduce evaporation and save you money!

        QPCR and PCR are common research focuses in many labs.  Many of us have experienced inconsistencies between samples or even triplicates from run to run or well to well.    Did you know that evaporation may be to blame?  A small change in the volume of buffers can alter the pH of the mix in the well, impacting the ability of the enzyme to function efficiently.  For instance, at 74C for every 5ul evaporation loss of a 25ul reaction, the pH changes by 1.   What causes this evaporation and how can we minimize it to result in more consistent, repeatable experiments?  Can we minimize evaporation so much that we can even downscale our reactions?

                A standard one-piece plate with a polypropylene frame expands by up to 2mm during thermal cycling (Figure 1).  The issues arise when the sealing film or strip caps expand and contract at different rates, leaving a small space and opening up the well to evaporation.  In addition, PCR blocks do not support PCR plates from the side, allowing for the high temperatures of the thermal block and heated lid to accelerate the expansion of the plates (Figure 2).

Evaporation in itself is an issue, but a large issue also arises when uneven evaporation is considered.  Generally evaporation occurs in the outside wells (Figure 3).  More evaporation in one well over another means wells may have different pHs and thus differences in enzyme efficiency.  This becomes an issue when comparing sample to sample or when trying to repeat experiments.

                Two-piece construction plates combat the issues of evaporation.  In two-piece construction plates, the tubes are made of polypropylene and the frame is polycarbonate.  Polycarbonate is a more rigid plastic that decreases the amount of thermal expansion, resulting in reduced evaporation and this improved PCR performance.  Two-piece construction plates, due to the lower evaporation, allow for downscaling of PCR reactions. 

                In comparing the volume loss of a one-piece plate to a two-piece construction plate, the researcher is losing money as each microliter of enzyme evaporates.  For instance, as seen in Table 1, the outer wells of a one-piece construction plate lose almost 3 µl while the inner wells lose over 1.5 µl due to evaporation.  4titude®’s Framestar™ two-piece construction plates, on the other hand, only lost 0.57 µl per well in the outer wells and a little over 0.5ul in the inner wells.  A loss of more than 6 times in the one-piece plates, adds up significantly in money loss.  If you were to assume the master mix costs about an average of 10 cents per microliter, then in the One-piece plates, a researcher loses $23.32 per each 96 well plate due to evaporation (Table 1).   In contrast, a researcher would only lose $5.33 in 4titude®’s Framestar™ two-piece construction plates.  In PCR plates, small amounts of evaporation per well result in not only reproducibility issues, but also in money loss.  Two-piece construction plates greatly reduce those issues.

               

              4titude's lower evaporation per well, a researcher can save money by reducing the reaction reagent volumes without compromising results.  Dr. Carolyn Denier in Berlin was able to reduce Sybr green usage by 40%, saving over $30 per plate and still maintain the consistent CTs she had in the past.  384 well users also see great results with 4titude® plates.  Dr. Andreas Dahl in Berlin was able to downscale his reactions as well due to decreased evaporation.  By reducing reaction volumes, a researcher can save significantly over time. 

                Overall, 4titude®’s Framestar™ plates and strip tubes have less thermal expansion for improved sealing integrity resulting in lower sample evaporation.  The lower sample evaporation allows customers to maintain more consistent results and downscale their reactions to save money.  Due to the lower thermal expansion of the 4titude® PCR plastics from MIDSCI, these plates are great for robotic use in that they are more reliable for pipetting and stacking.  The lower sample evaporation of 4titude’s PCR plastics makes these plates and strip tubes great for low volume PCR.  The plates also come in white well format for higher sensitivity and even greater consistency from well to well. 

Call MIDSCI today to purchase the 4titude® cost saving products!

Improve DNA collection, storage and purity with Isohelix Buccal Swabs and kits

                Buccal swabs are used across a wide variety of fields.  They are used in research laboratories, forensic labs, genetic testing laboratories, veterinary genotyping and diagnostics laboratories, paternity testing laboratories, and clinical diagnostics, to name a few.  The type of buccal swab used can not only have an effect on the amount of DNA collected, but can also impact further stability and extraction in downstream applications.  Isohelix swabs have a novel matrix that ensures efficient collection and increases DNA yield and ease of extraction.  When paired with Isohelix kits, even higher yields and purity can be obtained.

Isohelix provides an integrated solution for DNA processing with products for sampling, transporting, stabilizing, and extraction while maintaining high yields and purity.  Isohelix swabs provide significant advantages over other commercially available swabs. (Figure below)

What sets Isohelix apart?

•       High yielding swabs for buccal cell DNA collection

•       Unique swab matrix greatly improves DNA yields

•       Alternative to blood collection

•       Suitable for human and veterinary use

There are various different swab types available to fit your needs.  All swabs are ethylene oxide treated and are tested for DNA contamination.  The SK-1 swabs come with a 5 mL tube. After collecting the shaft can be easily snapped above swab head.  The SK-1 swabs are best paired with Dri-capsules, which allow for storage of the DNA for at least 3.5 years at room temperature!

The swabs can also be purchased with a 2 mL tube.  After collection, the lid on the 2 mL tube is securely closed.  If air venting is desired for drying of the swab in the field, the stopper can be opened while still having a closed cap.  The swab can then be removed by pulling on the shaft. 

If desired, swabs can also be purchased individually wrapped, two per pack, or three per pack. 

Need to effectively stabilize and/or isolate your DNA?

The DDK kit is a no column quick processing kit.  The DDK kit includes a stabilization reagent (stabilizes for up to 5 years) and isolation reagent that maintains both high yield and purity.  Up to 10 µg of DNA can be obtained from each swab with the DDK kit. 

The two cartridge based kits, XtraClean and Xtreme are important when extra high purity is desirable.  These kits are great when downstream analysis assays are arrays or quantitative PCR. 

Need your DNA Rapidly?

Try the BEK Buccalyse DNA Release Kit.  Obtain a 2 to 4µg yield in minutes! 

Isohelix swabs and kits can increase not only your DNA yield, but stability, purity, and ease of extraction.  Contact MIDSCI today for more information.   

Bioreactors: What are they and how can they improve experiments?

Have you ever had to clean out Erlenmeyer flasks or glass bottles after growing bacteria or suspension cells? Did you ever forget to autoclave bottles and then had to wait to start your experiment? Have you ever wanted to grow and spin suspension cells in the same vessel?  Would you like to vary gas exchange without opening up the culture to contamination?

Disposable bioreactors are the solution to many of these common issues in growing suspension and microbial cultures.  

What is a bioreactor? 

Bioreactors are growth vessels (of varying types) that provide similar cell physiology to in vivo allowing for optimal cell growth and high level productivity.

How do bioreactors accomplish this?

Bioreactors optimize cell growth and lead to high level productivity by maintaining the following characteristics:

•  Stable temperature
•  Maintenance of oxygen and gas level
•  Physiological pH
•  Low shear stress
•  Rapid response to induced changes

How to choose a bioreactor for your needs—(what are important qualities of a bioreactor?):

·        

• Ease of use
•  Rapid set up
•  Disposable
•  Efficient gas exchange and even dispersing of nutrients (Figure 1)
•  Low shear stress
•  Minimal impact of errors and contamination
•  Lower chance of contamination

            MIDSCI is the United States’ exclusive importer of TPP bioreactors (Figure 2). 

What sets TPP bioreactors apart?

•          TPP bioreactors have more efficient shaking (due to the internal cyclonic stirring) compared to the Erlenmeyer flask and bottle growth vessel (Figure3)

•          TPP bioreactors have no excess CO₂ as the efficient shaking allows for proper gas exchange and dispersal of nutrients (Figure 1)
•          TPP bioreactors have no excess foam or shear stress (Figure 4)

•          TPP bioreactors allow for customizable gas exchange with their 5 cap openings.  (Figure 5)

·        

• TPP bioreactors can be shaken and grown in a normal CO₂ incubator.
• 15 and 50 mL TPP bioreactors can easily be spun down in a centrifuge without transferring.  Rotors for the 600 mL TPP bioreactor are soon to come.  The less steps, the less of a chance there is of the introduction of contaminants. 
• Can easily scale up or scale down experiments.

MIDSCI has 15, 50, and 600 mL disposable bioreactors available in various packaging sizes.  The 15 mL is available in 200, 400, and 800 items/case.  The 50 mL is available in 60 and 180 items/case.  The 600 mL is available in 3, 13, and 26 items/case. 

Have any questions?  Interested in learning more about our bioreactors?  Contact MIDSCI today!