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FERTIUP® and CARD MEDIUM

FAQs
General questions regarding our products
Q1.Please tell me the storage method and storage period for FERTIUP® Mouse Sperm Cryoprotectant/
Mouse Sperm Preincubation Medium and CARD MEDIUM.
A1.

1. FERTIUP® Mouse Sperm Preincubation Medium and CARD MEDIUM should be stored at a temperature between 2~10℃.
2. FERTIUP® Mouse Cryoprotectant should be stored at a temperature between 18~28℃.
3. For the retention period, please see the documentation sent with the product, or alternatively the notes on the side of the ampoule.

Q2.Once opened, is it possible to divide a product into smaller amounts and cryopreserve it?
A2. No, it is not. We cannot guarantee the quality of our products after cryopreservation,
and so we do not recommend cryopreserving them.
Q3. Are there any substances contained in your products which may be harmful to humans?
A3. No, there are no substances in our products which are classed as harmful to humans by the Material Safety Data Sheet (MSDS).
Q4.How can I purchase reagent products produced by Kyudo Co. Ltd?
A4. 1. For orders within Japan, please contact us directly by e-mail. Our address is fertiup@kyudo.co.jp.
2. For orders from overseas, please contact our overseas supplier, Cosmobio Co., Ltd.
You can contact them using via e-mail, or alternatively you can visit their website.
 HP : Cosmobio Co., Ltd.
 E-mail : export@cosmobio.co.jp
Q5. How many days does it take for products to arrive?
A5. omestic orders are dependent on stock levels, but in principle we dispatch products via next-day delivery on the day after we receive an order.
This means that your products will arrive two days after we have confirmed your order.
For orders from overseas, please contact Cosmobio Co., Ltd.
Q6. There is a foreign object in the product. Can I receive a replacement product?
A6. Using our quality control system, we aim for the highest quality products from manufacture to shipping. In the unlikely event that you find glass fragments or other such foreign objects in a product,
please contact us and we will arrange for a replacement product to be sent to you.
Q7. There was precipitant in un-opened FERTIUP® Mouse Cryoprotectant.
A7. In case the precipitant is obtained, please incubate FERTIUP® Mouse Cryoprotectant ample 30 min to 1 hour at 37℃.After dissolving, there is no problem in use.
Questions regarding the FERTIUP® series of products
Q1. ou stated that you were able to obtain a high fertility rate for the C57BL/6 strain of mouse via in vitro fertilization using cryopreserved spermatozoa treated with FERTIUP® Mouse Cryoprotectant/Mouse Preincubation Medium.
Do the media increase the in vitro fertility rate of other strains too?
A1. We are currently investigating this further,
so please wait a little longer until we have compiled the relevant data.
Q2. was not able to use the entire 0.5mL of the FERTIUP® Mouse Cryoprotectant.
Would it be possible to use the cryoprotectant for a few weeks
if I were to transfer the medium into tubes and store it at refrigerated temperatures?
A2. No, it is not possible. Once open, we recommend using up an ampoule as quickly as you can do the following.
To do so, please purchase an ampoule of reagent which you can use in one occasion.
Please note that the ampoule of cryoprotectant should be stored at room temperature.
This is because the cryoprotectant contain high doses of saccharides and skim milk, which may sink to the bottom of the ampoule after refrigerating or freezing.
Q3. My FERTIUP® Mouse Sperm Cryoprotectant has turned white during its warrantee period.
Can I still use it?
A3. Yes, you can. The milky color is caused by casein micelle, a protein found in the skim milk in the cryoprotectant, settling and depositing on the bottom of the ampoule.
Casein micelle has no effect on the cryopreservation of sperm and so all efforts are made to remove as much of it as possible during manufacture, but occasionally a trace amount of casein micelle deposits at the bottom of the ampoule during long-term storage.
In the event that the casein micelle gets stirred into the product via tapping the ampoule, etc., the cryoprotectant appears milky white but has no adverse effect on the in vitro fertility rate.
Q4. Liquid paraffin is used in the usage instructions for the FERTIUP® series; does the paraffin have an adverse effect on the sperm?
A4. Yes, it does. However, as a rule liquid paraffin does not mix in with the sperm suspension (sperm suspended in preservant), so there is no adverse effect on the sperm.
Q5.

After preincubating cryopreserved sperm using FERTIUP® Mouse Preincubation Medium,
what is the sperm concentration when carrying out in vitro fertilization using an IVF medium?

A5. Normally, the sperm concentration in cryopreserved sperm ranges from 100,000~200,000 sperm/μL.
When introducing freeze-thawed sperm suspension (10μL) into our Mouse Sperm Preincubation Medium (90μL), the sperm concentration is diluted tenfold.
This means that the sperm concentration in the preincubation medium is 10,000~20,000 sperm/μL.

Thereafter, the preincubated sperm suspension (10μL) is further diluted tenfold when it is introduced into IVF medium (90μL).
This results in a sperm concentration in the IVF medium of 1,000~2,000 sperm/μL.

Finally, the ratio of motile sperm in after freeze-thawing is around 20%, which means that the motile sperm concentration in the IVF medium is 200~400 sperm/μL.
Q6. I would like to download the instruction manuals for the FERTIUP® series.
Where can I download them from?
A6. They are available on our website,  kyudo Co., Ltd.
Q7. Is there a method or measure to confirm whether the freezing and thawing of sperm has been correctly carried out?
A7. 1. When simply confirming the quality of the sperm after freeze-thawing, preincubate the sperm in preincubation medium after freezing and judge according to the number of active motile sperm at the edge of the drop.
2. When carrying out final confirmation, carry out in vitro fertilization of unfertilized oocytes and then confirm that you have attained at least a 30% fertility rate.

(We carry out IVF during lot checks to check and confirm that all of our products on sale attain an in vitro fertility rate of 30% or above for C5BL/6 mice)
Q8. How about legacy stock of cryopreserved spermatozoa?
A8. Please Check here. Rescue In Vitro Fertilization Method for Legacy Stock of Cryopreserved Spermatozoa
uestions regarding CARD MEDIUM
Q1. For how many months after manufacture does the warrantee period for CARD MEDIUM last?
A1. CARD MEDIUM is a new product and so at present, the warrantee period is 18 months. This warrantee period may increase in the future, depending on the results of investigations and reviews.
Q2. Is it possible to attain a high in vitro fertility rate without using CARD MEDIUM in association with the FERTIUP® series?
A2. No, it is not. The FERTIUP® series works on sperm and CARD MEDIUM works on oocytes in order to raise the fertility rate. As such, using both products simultaneously is the key to attaining a fertility rate of over 80% even for the C57BL/6 strain of mouse.
Q3. Are we able to attain a high in vitro fertility rate for strains apart from the C57BL/6 strain of mouse?
A3. Yes, other mouse strains also show high in vitro fertility levels.