SAMPLING PROTOCOL

STANDARD AQUATIC eDNA KIT

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USING THE KIT

KIT CONTENTS

Kit contents are based on one person collecting an eDNA sample. If two or more people will be handling kit contents and collecting an eDNA sample, please contact us about inclusion of extra gloves in kits. Extra gloves are not provided as standard to minimise waste.

1x pair of nitrile gloves

1 x 3.5 L sampling bag
1 x specimen bag

1 x enclosed filter (0.8 μm pore size, polyether-sulfone)

1 x 100 mL Luer Lock plastic syringe

1 x sampling datasheet

1 x resealable bag containing 1 x small syringe filled with 1.5 mL of DNA preservative solution (sealed with reusable Luer Lock cap) and 1 x separate Luer Lock cap

EXPERIENCED USER PROTOCOL
DETAILED USER PROTOCOL

EXPERIENCED USER PROTOCOL

1. Put on the provided gloves. Use the provided sampling bag or a new mineral water bottle (with the water discarded) to collect your (sub)samples from the waterbody (see Appendix). Minimise contact with the water during collection. Deposit the (sub)sample in the sampling bag, seal, and make sure the water is well mixed by shaking for 20-30 seconds. You should use a new bag/bottle for each sample (i.e. each kit) and for each waterbody to avoid cross-contamination.

2. Draw up 100 mL of water from the sampling bag into the large syringe. Attach the syringe to the filter inlet (narrow end). Press the plunger to push the water through the filter.

3. Repeat steps 1 and 2 until all water has been filtered or the filter clogs. Make a note on the sampling datasheet of the total volume processed.

4. Detach the syringe from the filter, hold the filter in one hand, and pull back the plunger to fill the syringe with air. Reattach the filter and push the air through to expel any water trapped inside the filter. Repeat several times to remove as much water as possible.

5. Uncap the small syringe (already filled with preservative solution) and twist it onto the filter inlet. Do not discard the Luer Lock cap – hold the cap in one hand as it will be needed in step 6.

6. Hold the filter so that the outlet (wide end) points upwards. Carefully and slowly press the plunger to push the preservative solution into the filter. Stop when the first drop can be seen emerging from the filter outlet, but do not remove the small syringe with preservative solution. Cap the filter outlet using the Luer Lock cap that was on the small syringe in step 5.

7. Invert the filter so that the filter outlet points down, and slowly press the plunger to expel the rest of the preservative solution. The entire volume of preservative solution should be added to the filter and the small syringe should be empty. Detach the small syringe whilst keeping the plunger depressed and cap the filter inlet with the separate Luer Lock cap.

8. Place the filter inside the specimen bag and seal. If storing samples at ambient temperatures, samples should be received by the lab within 2 weeks of sampling. If this is not possible, we recommend that samples are frozen as soon as possible after sampling to enhance DNA preservation until samples are sent back to us. Cold shipping is not necessary for frozen samples. Please contact NatureMetrics if return within 2 weeks or long-term freezing is not possible.

9. Complete the sampling datasheet and use the NatureMetrics app to scan the barcode on the sampling datasheet to complete your online submission.

10. Place the bagged filter and sampling datasheet in the bag that the kit contents arrived in. For UK sample returns, simply use the standard postal service with recorded delivery. Follow the packaging notes and send to: Nature Metrics, 1 Occam Court, Surrey Research Park, Guildford, GU2 7HJ, United Kingdom. For international sample returns, contact your NatureMetrics Business Development Manager or our Logistics team (logistics@naturemetrics.co.uk) for further information. We are only able to analyse samples that are returned using the correct NatureMetrics logistics procedure. 

DETAILED PROTOCOL

PART 1: COLLECTING THE WATER

Ideal sampling strategies will vary among habitats and monitoring contexts. We recommend that you speak to a member of our team to plan your survey. We also recommend the inclusion of field negative controls (blanks) to detect possible contamination introduced during sampling. NatureMetrics provide a free kit to be used as a field blank if 20 or more Standard Aquatic eDNA Kits are purchased.

Download and log into the NatureMetrics app before arriving in the field. Avoid introducing DNA (from yourself, soil or other waterbodies) into the water and prevent spread of disease or invasive species by wearing the provided gloves and minimising contact with the water.

PART 2: FILTERING THE WATER AND PRESERVING THE DNA

1. Draw up 100 mL of water from the sampling bag into the large syringe. If there is a high sediment load, then leave the sample for a minute to allow the sediment to settle before drawing water up into the syringe. This should increase the amount of water that you can filter.

2. Attach the syringe to the filter inlet (narrow end). The syringe should easily twist onto the inlet side of the filter. Press the plunger to push the water through the filter.

3. Repeat steps 1 and 2 (holding the filter in one hand while it is not attached to the syringe) until all water has been filtered or the filter clogs. Keep track of the number of times the syringe is refilled. Make a note on the sampling datasheet of the total volume processed.

4. Detach the syringe from the filter, hold the filter in one hand, and pull back the plunger to fill the syringe with air. Reattach the filter and push the air through to expel any water trapped inside the filter. Repeat several times to remove as much water as possible. It helps to gently shake the filter as you do this.

PART 3: PRESERVING THE DNA

1. Uncap the small syringe (already filled with preservative solution) and twist it on to the filter inlet. Do not discard the Luer Lock cap – hold the cap in one hand as it will be needed in step 2. The solution is non-hazardous to aquatic life but release into the environment should be avoided. We advise performing this step away from the water’s edge and avoiding spills.

2. Hold the filter so that the outlet (wide end) points upwards. We advise you to do this step away from anyone’s eyes. Carefully and slowly press the plunger to push the preservative solution into the filter. This must be done slowly to allow the preservative solution to spread out over the filter surface. Stop when the first drop can be seen emerging from the filter outlet, but do not remove the small syringe with preservative solution. Cap the filter outlet using the Luer Lock cap that was on the small syringe in step 1.

3. Invert the filter so that the filter outlet points down, and slowly press the plunger to expel the rest of the preservative solution. The entire volume of preservative solution should be added to the filter and the small syringe should be empty. Detach the small syringe whilst keeping the plunger depressed and cap the filter inlet with the separate Luer Lock cap. The preservative solution contains a detergent – don’t be alarmed if it foams a little!

4. Place the filter inside the specimen bag and seal. If storing samples at ambient temperatures, samples should be received by
the lab within 2 weeks of sampling. If this is not possible, we recommend that samples are frozen as soon as possible after sampling to enhance DNA preservation until samples are sent back to us. Cold shipping is not necessary for frozen samples. It should be noted that although freezing is recommended as best practice for optimal storage, we understand that it is not always feasible. Storing at ambient temperatures is a tried and tested method that yields excellent data. The addition of freezing is to further safeguard the samples against potential DNA degradation, which can affect species detection when samples are stored at high temperatures for long periods. Please contact NatureMetrics if return within 2 weeks or long-term freezing is not possible.

You can view a video demonstration of filtration with the Standard Aquatic eDNA Kit here.

PART 4: RETURNING YOUR SAMPLES

Once you have collected and filtered your sample, follow our guidelines below to safely transport your sample to the NatureMetrics laboratory.

1. Complete the sampling datasheet and use the NatureMetrics app to scan the barcode on the sampling datasheet to complete your online submission. Please record your sampling data on paper and through the app so there is a physical and electronic copy of your data.

2. Place the bagged filters and sampling datasheet inside the bag that the kit contents arrived in.

3. For UK sample returns, simply use the standard postal service with recorded delivery. Follow the packaging notes and send to: Nature Metrics, 1 Occam Court, Surrey Research Park, Guildford, GU2 7HJ, United Kingdom.

3. For international sample returns, contact your NatureMetrics Business Development Manager or our Logistics team (logistics@naturemetrics.co.uk) for further information. NatureMetrics is fully compliant with current legislation on the transport of biological material and our operations team are on hand to ensure that all return shipments meet the required specification. We are only able to analyse samples that are returned using the correct NatureMetrics logistics procedure. 

PACKAGING NOTES AND QUICK LINKS

  • Return your samples using the box that kits were sent in, where possible.

  • If using an alternative box, ensure that it is in good condition and remove any old labels.

  • Avoid excess spacing by adding cushioning material (e.g. bubble wrap, newspaper) to avoid the product shifting inside the box.

  • Use good quality sealing tape and seal along box edges in a H pattern.

  • The only essential thing to send back to us is the specimen bag containing the filter and the sampling datasheet. However, all kit contents can be sent back to NatureMetrics for recycling.

  • Register for your my.naturemetrics account here.

  • Please find our Terms and Conditions here.

CLIENT RESPONSIBILITIES

Important considerations for your results

  • The absence of a detection does not necessarily imply the absence of a taxon from a location.

  • Samples from some locations might have compounds that inhibit analyses resulting in reduced data generation – while we routinely test for inhibition, it is not always possible to overcome.

  • It is the responsibility of the client to ensure that all efforts have been made to avoid contamination from external sources and between samples.

  • Handling samples without gloves can increase the content of human DNA, reducing data generation.

  • After sampling, movement of samples across borders without permission is not allowed as they are classed as biological samples.

Important considerations for interpreting your results

It should be noted that DNA can enter an ecosystem via many routes (e.g. wastewater from commercial and domestic sources). While DNA from a given taxon may be present and detectable, it is not possible to discern the source of the DNA. Results from common food items, domestic species and livestock species need to be interpreted with caution.

Disclaimer: Safe sample collection is the responsibility of the Client. NatureMetrics accepts no liability associated with the use of the kits and sample collection. The Client is solely responsible for the quality of the samples and the representativeness of the samples received by NatureMetrics. The information contained within the Final Report provided by NatureMetrics to the Client is not intended to be advisory, it is informational. Interpretation and decisions are the sole responsibility of the Client. NatureMetrics does not accept any liability whatsoever for any reliance placed on any information contained within, or any use that may be made of, the Final Report by the Client. Please read the full limitation of liability statement in the Terms and Conditions.

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