Sato Laboratory, Department of Health Sciences (Neurology), Graduate School of Biomedical Sciences, Nagasaki University

Prion Disease-Related Test (ENGLISH)

JAPANESE

We are conducting cerebrospinal fluid (CSF) testing for human prion diseases in cooperation with and on behalf of the "Prion Disease and Delayed Viral Infection Research Group (Takao Group)" and the "Creutzfeldt-Jakob Disease (CJD) Surveillance Committee (chaired by Yamada and Mizusawa)" under the project to overcome intractable diseases funded by the Ministry of Health, Labor and Welfare through the Chemical Research Grant. The testing is being performed by the CJD Surveillance Committee (ELISA for 14-3-3 protein and total tau protein in the CSF is available free of charge).
We have also succeeded in detecting a small amount of abnormal prion proteins in the CSF and established a real-time quaking-induced conversion (RT-QuIC) assay. Please make a separate request if you wish to undergo the test. The test will be performed free of charge initially, but we may ask you to pay a part of the cost in the future. For urgent (within one week) testing (e.g., to rule out prion disease prior to brain biopsy), please contact us separately. 14-3-3 protein, total tau protein, and RT-QuIC detection of abnormal prion proteins can be performed within one week (Please inquire at

nagasakiprion@yahoo.co.jp).

Please see each item for more details:

1) CSF testing (total tau and 14-3-3 proteins)
2) Research and establishment of biomarkers for rapidly progressive dementia including prion diseases
3) Urgent cases where the results are needed within one week
4) Q&A

Notice:

April 18, 2023
Notice of Inspection Holiday
We will accept urgent cases until April 24.
Normal business will begin on May 8.
We apologize for any inconvenience.

	New Dr. Atarashi's paper has been published in February 2011 Volume 17, Issue 2 NATURE MEDICINE

Information on Prion disease-related test:

	Test Request Form（MS-Word 13KB）
	Requests for Test Request Form（MS-Word 16KB）
	Checkist before sending specimens（MS-Word 14KB）
	Study Protocol（MS-Word 325KB）

Checklist before sending samples:

・	We do not inform the test results by phone.
・	Although the tests are performed by the Department of Neurology, Nagasaki University Graduate School of Biomedical Sciences, the specimens should be sent to the former Nagasaki University School of Medicine, 1st Department of Anatomy. Please do not make any errors.
・	Please refer to"Requests for Specimens"for information on how to transport the specimens.

1) CSF testing (14-3-3 and total tau protein testing)
We are currently accepting subcontracted testing for total tau protein at SRL for reimbursement. We would be honored if you could use the results.

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2) Research and establishment of biomarkers for rapidly progressive dementia, including prion diseases
Currently, the diagnosis of rapidly progressive dementia, including prion diseases, is made using a combination of MRI diffusion-weighted imaging, spinal fluid testing, electroencephalography, and genetic testing, in addition to clinical symptoms. However, these tests are only adjunctive. A definitive diagnosis of rapidly progressive dementia can only be confirmed by the detection of abnormal proteins in the brain tissue and immunohistochemistry after postmortem brain biopsy or pathological autopsy. Therefore, the only method to confirm the diagnosis before death with a possibility for treatment is a highly invasive brain biopsy. Therefore, we aim to identify novel related substances to establish a diagnostic method that can confirm dementia before death and predict prognosis. We will identify "substances related to rapidly progressive dementia" using biological materials collected from living patients diagnosed with rapidly progressive dementia by the above-mentioned auxiliary tests, and establish a biomarker and protein detection method for these substances. The RT-QuIC assay is an amplification method for abnormal prion proteins that can detect ultra-trace amounts of abnormal prion proteins in the brain tissue or CSF. This method is equivalent to the polymerase chain reaction method for proteins. The RT-QuIC assay (Nat Methods. 2008 Mar;5(3):211-2) was developed by our laboratory and can detect solitary Creutzfeldt-Jakob disease with a sensitivity of 90-95% and a specificity of almost 100%. We are also measuring 14-3-3 and total tau protein levels in the CSF of patients with prion diseases.

2) Research and establishment of biomarkers for rapidly progressive dementia, including prion diseases

Currently, the diagnosis of rapidly progressive dementia, including prion diseases, is made using a combination of MRI diffusion-weighted imaging, spinal fluid testing, electroencephalography, and genetic testing, in addition to clinical symptoms. However, these tests are only adjunctive. A definitive diagnosis of rapidly progressive dementia can only be confirmed by the detection of abnormal proteins in the brain tissue and immunohistochemistry after postmortem brain biopsy or pathological autopsy. Therefore, the only method to confirm the diagnosis before death with a possibility for treatment is a highly invasive brain biopsy.
Therefore, we aim to identify novel related substances to establish a diagnostic method that can confirm dementia before death and predict prognosis. We will identify "substances related to rapidly progressive dementia" using biological materials collected from living patients diagnosed with rapidly progressive dementia by the above-mentioned auxiliary tests, and establish a biomarker and protein detection method for these substances.

The RT-QuIC assay is an amplification method for abnormal prion proteins that can detect ultra-trace amounts of abnormal prion proteins in the brain tissue or CSF. This method is equivalent to the polymerase chain reaction method for proteins. The RT-QuIC assay (Nat Methods. 2008 Mar;5(3):211-2) was developed by our laboratory and can detect solitary Creutzfeldt-Jakob disease with a sensitivity of 90-95% and a specificity of almost 100%. We are also measuring 14-3-3 and total tau protein levels in the CSF of patients with prion diseases.

＊	The RT-QuIC assay is not a 100% accurate test. False-positive and false-negative results are possible. Please make a comprehensive judgment based on the clinical course and MRI images. The sensitivity and specificity of the RT-QuIC assay vary depending on the clinical course of the disease. Therefore, we would be grateful if you could re-submit a case that was biomarker-negative in the early stage of disease onset.

＊

The RT-QuIC assay is not a 100% accurate test. False-positive and false-negative results are possible.
Please make a comprehensive judgment based on the clinical course and MRI images.

The sensitivity and specificity of the RT-QuIC assay vary depending on the clinical course of the disease. Therefore, we would be grateful if you could re-submit a case that was biomarker-negative in the early stage of disease onset.

Sample
Send plasma and CSF frozen in screw-cap tubes for cryopreservation. (If you have a family member who agrees to this, please send us the specimen at the same time.)
Application Forms

Process

1. Application by e-mail at nagasakiprion@yahoo.co.jp

2. Acceptance: A procedure number is issued by e-mail from our office.

Download the below Surveillance Committee Questionnaire
from the website and send it to the applicant via e-mail.
Email the Excel file to Nagasaki University (

nagasakiprion@yahoo.co.jp
and to Nagasaki University (

prion-ncnp@ncnp.go.jp) on CC.

3. Sending the specimens

*Transportation should be frozen, and "Receipt No. 0000" should be clearly indicated on the addressed item.

*Please send to the following address:

3F, Basic Building, 1-12-4 Sakamoto, Nagasaki 852-8523, Japan Nagasaki University School of Medicine Former First Dissection Katsuya Satoh

4. Emailing of the results

	Surveillance Survey Form（MS-Excel 231KB）
	Research and Establishment of Biomarkers for Rapidly Progressive Dementia including Prion Disease Explanation and Consent Form for the Research on "Biomarkers of Rapidly Progressive Dementia Including Prion Disease"（MS-Word 66KB）
	Prion Disease Surveillance Committee（PDF 960KB）
	Explanatory Document, Consent Form, Consent Confirmation Form, Consent Withdrawal Form, Consent Confirmation Form

Sending and Shipping Address for Samples
Samples should be sent to the same address as for 14-3-3 protein and total tau protein.
Cost
Initially, no charge is applicable. A fee may be incurred in the future.
Precautions
It is known that the detection sensitivity is slightly reduced in cases of CSF containing more than 100 mg/dl of protein or a large amount of blood. CSF with as little blood as possible is desirable. CSF that has been repeatedly frozen and thawed decreases the detection sensitivity. Therefore, it is preferable to submit CSF samples that have not been frozen and thawed as much as possible. The RT-QuIC assay may yield false positive results in patients with epilepsy. Please use caution during the determination.
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3) Urgent cases where results are needed within one week:
In cases where results are urgently needed within a week, such as cases of rapidly progressive dementia, malignant lymphoma, intracerebral malignant lymphoma, and other cases where brain biopsy is performed and prion disease cannot be ruled out on imaging, we offer a service to detect 14-3-3 protein in the CSF, total tau protein, and abnormal prion protein using the RT-QuIC assay within a week. We would be honored if you could send us an email describing the urgency of your request. Please be sure to include an abstract of your specimen request. We do not issue quotations.

3) Urgent cases where results are needed within one week:

In cases where results are urgently needed within a week, such as cases of rapidly progressive dementia, malignant lymphoma, intracerebral malignant lymphoma, and other cases where brain biopsy is performed and prion disease cannot be ruled out on imaging, we offer a service to detect 14-3-3 protein in the CSF, total tau protein, and abnormal prion protein using the RT-QuIC assay within a week.
We would be honored if you could send us an email describing the urgency of your request.
Please be sure to include an abstract of your specimen request.
We do not issue quotations.

Specimens, Application Forms, Sample Delivery and Mailing Address:
The same as for 14-3-3 protein, total tau protein, and the RT-QuIC assay. You are responsible for your own expenses. You will be invoiced for the cost by the Accounting Section of the Finance Department of Nagasaki University.
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Request for sending images of patients with suspected prion disease:
As part of the activities of the CJD Surveillance Committee, we have conducted a "Reading Study on the Usefulness of Standardized Diffusion-Weighted Images in Determining Disease in Creuzfeldt-Jakob Disease" with your cooperation, and have suggested the usefulness of standardizing the display conditions of diffusion-weighted images. It is important to establish the diagnostic accuracy of these images by conducting prospective studies. Therefore, we would appreciate it if you could send us MRI images (diffusion-weighted images, b0 images, ADC maps, and FLAIR) in DICOM format when you request a CSF examination. If you have performed cerebral blood flow studies, we would appreciate it if you could send us the images as well. Additionally, we would appreciate it if you could delete the MRA files. Please save the images on a CD-R or other media and send them to us. If possible, we would appreciate it if you could anonymize the header information, such as the patient's name and age, so that the individual cannot be identified. The images you send us will be managed by Masashi Harada, Department of Radiology, University of Tokushima Hospital (in charge of imaging by the Prion Surveillance Committee), and will be used for surveys and research as part of the surveillance. We apologize for any inconvenience this may cause in your busy schedule, but we sincerely appreciate your cooperation.

Request for sending images of patients with suspected prion disease:

As part of the activities of the CJD Surveillance Committee, we have conducted a "Reading Study on the Usefulness of Standardized Diffusion-Weighted Images in Determining Disease in Creuzfeldt-Jakob Disease" with your cooperation, and have suggested the usefulness of standardizing the display conditions of diffusion-weighted images.

It is important to establish the diagnostic accuracy of these images by conducting prospective studies. Therefore, we would appreciate it if you could send us MRI images (diffusion-weighted images, b0 images, ADC maps, and FLAIR) in DICOM format when you request a CSF examination. If you have performed cerebral blood flow studies, we would appreciate it if you could send us the images as well.
Additionally, we would appreciate it if you could delete the MRA files.

Please save the images on a CD-R or other media and send them to us. If possible, we would appreciate it if you could anonymize the header information, such as the patient's name and age, so that the individual cannot be identified. The images you send us will be managed by Masashi Harada, Department of Radiology, University of Tokushima Hospital (in charge of imaging by the Prion Surveillance Committee), and will be used for surveys and research as part of the surveillance. We apologize for any inconvenience this may cause in your busy schedule, but we sincerely appreciate your cooperation.

Request for Cooperation in a Naturalistic Survey of Prion Disease Patients:
The Japanese Consortium of Prion Disease (JACOP) is being organized as part of the Prion Surveillance Committee. The natural history of patients with prion disease is important for future clinical trials. We would be honored if you would cooperate with us in our efforts to overcome prion diseases. JACOP (Japanese Consortium of Prion Disease)

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4) Q&A

	The address has been changed since March 2021 to the following: 3F, Basic Building, 1-12-4 Sakamoto, Nagasaki 852-8523, Japan Nagasaki University School of Medicine Formerly First Department of Anatomy Katsuya Sato
	When transporting specimens, the Nagasaki University School of Medicine used to perform molecular analysis of infection. Is the address still the same?
	Is there a specified shipping company for specimens?
	No, we do not have a specific shipping company.
	I cannot specify a time between 10:00 AM and 11:00 AM for transportation. What should I do?
	If you cannot specify the time, please specify a weekday morning.
	Do you provide a screw cap to send specimens? What should I do?
	Please use a tube with a screw cap that will not open, as a normal Eppendorf tube is likely to open even if it is wrapped in paraffin.
	Is temperature control necessary for specimen transport?
	Specimens should be transported at a temperature of -20°C. If this is not possible, please send specimens with large amounts of dry ice.
	How much spinal fluid is needed for the test?
	The minimum amount of CSF required for testing is 0.6 cc. The minimum serum volume required for the test is 0.6 cc.
	Inquiries about test results
	Inquiries about test results are not available via phone.
	How long does it take to get the test results back?
	It takes approximately 2-3 weeks for 14-3-3 protein and total tau protein results, and approximately 2 months for detection of abnormal prion protein in the CSF (RT-QuIC method). If you want to know the results as soon as possible, please request an expedited service. The results will be provided within one week of the arrival of the specimen. There is no charge for the regular test; however, a 25,000 yen fee is required for early test results.
	Can an individual patient apply for a test?
	Currently, we are not accepting individual applications. We are very sorry.
	Can the patient or the patient's family ask Nagasaki University directly about the test results?
	We are sorry. Currently, we are unable to provide the results to patients or their families directly.
	Please provide information about the pretreatment procedures required when collecting the plasma before sending it for testing.
	These are the most frequently asked questions. The steps are as follows: Draw blood for one plasma collection tube (about 10 ml of EDTA-2K) with a purple cap. Centrifuge at 1,500–2,000×g for 15 minutes. Once the plasma has been separated, place it in a sampling tube (Eppendorf tubes) and seal it so that it does not leak outside. From this plasma, we measure the plasma neurofilament light chain (NfL) and attempt to correlate it with prognosis. Thank you for your cooperation.

(1)	If a specimen is sent by a method not described in the specimen transport manual and a safety issue is recognized, the test may be suspended.
(2)	Please do not attach any personal information of the patient to the test request form.
(3)	Test results do not confirm the diagnosis of prion disease. Please make a comprehensive judgment based on the clinical course, MRI images, etc.
(4)	Please consult with us before presenting the test results at an academic conference or in a paper.
(5)	This test is conducted as part of the research activities of the Ministry of Health, Labor and Welfare. Anonymized information may be published in academic journals, conferences, etc. in a summarized form.

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