vitamin d_pi
Contents
1. Vitamin D Package Insert One cartridge contains Monoclonal mouse anti vitamin D 8 0 8 ng Monoclonal anti testosterone antibody 76 7 6 ng Fluorescent particles 4 8 0 48 ng One pretreatment tube contains Gold nano particle conjugation antibody 5 0 5 ug One dilution tube contains Perfluorohexanoic Acid 1 4 0 14 mg Materials required but not provided The FREND System Micro pipette capable of delivering 35 and 70 uL Heating block for tube and cartridge incubation at 37 C Timer for incubation step Personal protective equipment and biohazard waste Warning and Precautions The FREND Vitamin D cartridges are intended for in vitro diagnostic use only Vitamin D cartridges are only to be used on the FREND System Vitamin D cartridges are disposable single use devices Do not reuse them under any circumstances Allow sealed cartridges to come to room temperature for 15 30 minutes prior to use Reagent kit should not be frozen Assure the humidity in the laboratory is in the 10 80 range when tests are run Assure the room temperature remains in the range of 22 30 when tests are run Avoid cross contamination between samples by using a new pipette tip for each new specimen Avoid high humidity direct sunlight or heat in the area used for cartridge storage Inaccurate results are possible if the sample used is contaminated in any way Using specimens containing clotted fibrin could result in erro2. 0 7940 Fax 82 2 6220 7721 EC REP MT Promedt Consulting GmbH Altenhofstrasse 80 66386 St Ingbert Germany US Branch NanoEnTek USA Inc 5627 Stoneridge Drive Suite 304 Pleasanton CA 94588 USA Tel 1 925 225 0108 1 888 988 0108 Toll free Fax 1 925 225 0109 China Branch SK Medical Beijing Co Ltd 26F SK Tower No 6 Jia Jianguomenwai Avenue Chaoyang District Beijing 100022 P R China Tel 86 10 5920 7844 Fax 86 10 5920 5697 Revised on 2015 10 01
3. NESPI FRVD 001EN V 0 1 Nano ntTek FREND Vitamin D Total 25 Hydroxyvitamin D Intended use The FREND Vitamin D test is a fluorescent nanoparticle immunoassay designed for in vitro quantitative measurement of total 25 hydroxy OH vitamin D and related hydroxylated metabolites in human serum and plasma EDTA Lithium heparin and Citrate The FREND Vitamin D microfluidic flow cartridge is designed for use in the FREND System to aid in the assessment of vitamin D sufficiency Summary and explanation of test Vitamin D is a fat soluble prohormone known for its role in regulating calcium and phosphorus levels in bone mineralization Sunlight exposure produces vitamin D via photochemical conversion of 7 dehydrocholesterol in the epidermis and is the primary source of vitamin D Seasonal changes amount of exposure sunscreen use and skin pigmentation can cause variation in the amount of vitamin D produced in the body A minor source of vitamin D can be absorbed from food and vitamin supplements with an estimated 10 20 sa absorbed by the body in this manner In circulation 25 OH vitamin D is bound to vitamin D binding protein VDBP or albumin at 1000 times higher concentrations than the active form 1 25 OH 2 vitamin D Additionally the 25 OH form has a half life of 2 3 weeks as compared to the less stable 1 25 OH 2 form which has a half life of a few hours 25 OH vitamin D exists as D2 ergocalcifer
4. artridge contains built in control feature Fluorescence signal in the reference zone of each cartridge shows 1 that enough volume is added 2 that proper flow is obtained and 3 that the antibody is reactive If this reference zone signal is missing or lower than threshold the FREND System consider it as an incorrect or failed test not producing a test result but an error message In addition with each cartridge run the system monitors in part for 1 flow of sample 2 speed of sample flow 3 shelf life of cartridge components 4 function of internal barcode scanner and 5 function of scanner s mechanical components External quality control testing Commercially available controls from a variety of manufacturers are available that contain 25 hydroxy vitamin D as a measured analyte It is recommended that a minimum of two 2 levels of controls be run at least once per month or once for each new lot whichever comes earlier However Controls should be run with a minimum frequency depending on number of tests run in the laboratory Each laboratory should establish its own criteria based on the following parameters 1 Each new lot 2 Each new shipment even if from the same lot previously received 4 Monthly as a continued check on storage conditions 5 Whenever problems storage operator or other are identified 6 Or other times as required by your laboratory s standard QC procedures Individual labor
5. atory policy will dictate exactly which control materials and lot numbers should be run the frequency with which controls are to be tested criteria for acceptance of the results and required corrective action to be taken if results do not meet laboratory criteria If any external quality control sample values are out of the acceptable range it will be necessary to investigate the problem before reporting patient results to assure there is not an instrument or software malfunction Do not assay patient samples on the FREND System using FREND Vitamin D if quality control on how to determine acceptability of external control material results Each laboratory operates under a different set of regulations Every laboratory must follow the standardized procedures acceptable to the regulatory agencies to which the laboratory is responsible Specimen processing Preparation Remove sufficient cartridges and pretreatment tubes of FREND Vitamin D from the refrigerator to test the number of patient samples and required external quality materials Allow the tubes and the sealed pouches containing the cartridges to come to room temperature for 15 30 minutes prior to the start of the testing sequence Heating block provided with FREND System should be turned on 7 8 minutes before use If using refrigerated patient samples remove those from the refrigerator and allow to them to come to room temperature prior to testing If frozen samples will be u
6. e HAMA anti goat HAGA or anti rabbit HARA antibodies maybe show alsely elevated or depressed values or may result in an incomplete test 11 12 Patients routinely exposed to animals or animal serum products can be prone to these types of heterophilic interferences 4 Certain medications may interfere with assay performance All results should be interpreted with respect to the clinical picture of the patient 5 Although hemolysis has an insignificant effect on the assay hemolyzed samples may indicate mistreatment of a specimen prior to assay and results should be interpreted with caution 6 Lipemia has an insignificant effect on the assay except in the case of gross ipemia where interference with the lateral flow of the sample in the cartridge may occur 7 The concentration of Vitamin D in a given sample determined with assays rom different manufacturers can vary due to differences in assay methods calibration and antibody specificity Please refer to the Specimen Collection and Handling Warnings and Precautions Storage and Stability and Procedural Notes sections in this insert sheet 9 FREND Vitamin D has not been validated in point of care settings 10 FREND Vitamin D is to be used in licensed clinical laboratories with trained technologies 8 Performance characteristics Performance characteristics were evaluated for the FREND Vitamin D as follows 11 Precision Clinical and Labora
7. g the arrows 3 Press the Setup button on the Main screen 4 Press the Code chip button on the Setup screen 5 The information embedded on the FREND Vitamin D Code chip is automatically saved on the FREND System 6 When the Code chip installation is completed press the OK button to go to the Setup screen 7 Press the Item button on the Setup screen 8 Check the FREND Vitamin D cartridge lot number and the installation date of the Code chip 9 Press the Home button to go to the Main screen to begin running external quality control and patient samples Quality control FREND System QC cartridges FREND QC Cartridge contains multiple controls to check optic part of the system By testing QC Cartridge part of analytical components of the system of 1 laser power 2 alignment and 3 mechanical integrity are confirmed For each day of patient testing perform QC Cartridge testing Refer to the quality control procedures section in the User Manual of FREND System In brief perform QC Cartridge testing for the following conditions 1 Upon initial setup of the system 2 Each day of patient testing 3 When the system has been transported or moved 4 Whenever there is uncertainty about the performance of the system 5 Whenever required by your laboratory s quality control requirements Internal procedural controls FREND Vitamin D test c
8. neous results Over or under loading the cartridge with sample may result in inaccurate results Human specimens are not used in the preparation of this product however since human specimens will be used for samples and other quality control products in the lab may be derived from human materials Please use Universal Precautions when handling all specimens and controls Do not use the reagent components beyond the expiration date on the pouch Do not use the reagent components if the pouch is damaged or the seal is broken Perform testing as specified in the Package Insert and User Manual Keep the reagent components sealed in the pouch until just ready for use Use the reagent component immediately after opening the pouch Wear disposable gloves when handling the reagent components and the samples Wash hands thoroughly and often after handling reagent cartridges or samples Do not ingest the silica gel package found in the cartridge pouch Vitamin D has been designed so that the high dose hook effect does not affect the vast majority of samples Handle specimens in accordance with the OSHA Standard on Bloodborne na Pathogens Storage and Stability All unopened materials are stable until the expiration date on the label when stored at the specified temperature Reagent stability has been demonstrated for twelve months from the date of manufacture The expiration date is clearly indicated on the product box and the car
9. of human serum vitamin D binding protein Biochemistry 1979 18 8 1611 1617 8 Holick MF et al Vitamin D2 is as effective as vitamin D3 in maintaining circulating concentrations of 25 hydroxyvitamin D The Journal of Clinical Endocrinology and Metabolism 2008 93 3 677 681 9 Maxmen A Nutrition advice the vitamin D lemma Nature 2011 475 7354 23 25 10 Chao E L Henshaw J L Occupational Safety and Health Administration Model Plans and Programs for the OSHA Bloodborne Pathogens and Hazard Communications Standards OSHA 3186 06R 2003 11 Schroff R W Foon K A Beatty S M Oldham R K Morgan A C Human Anti Murine Immunoglobulin Responses in Patients Receiving Monoclonal Antibody Therapy Cancer Research 1985 45 879 885 12 Boscato L M Stuart M C Heterophilic Antibodies A Problem for All Immunoassays Clinical Chemistry 1988 34 1 27 33 15 Glossary of symbols Do not reuse 2 Ep OVYVEMNDO Use by YYYY MM DD Lot number Catalog number A Warning or Caution Manufactured by EC REP Authorized representative in the Europe Community Invitro diagnostic medical device zc Temperature limitation Date sc V n Contains sufficient for lt n gt tests 16 Nano nfek sales nanoentek com www nanoentekinc com Manufactured by NanoEnTek Inc HQ 12F 5 Digital ro 26 gil Guro gu Seoul 08389 Korea Tel 82 2 622
10. ol and D3 cholecalciferol isomers with supplements available for both Often total 25 OH vitamin D is measured to assess the sufficiency in a patient and make appropriate clinical decision Principle of the assay The FREND Vitamin D test cartridge is a one time use rapid competitive immunoassay utilizing fluorescent nanoparticle in microfluidic flow to capture and quantify total 25 OH vitamin D in serum and plasma specimens The drop of 70 uL patient sample is placed in the FREND Vitamin D pretreatment tube where the sample interacts with a proprietary mix of pretreatment solution Initially patient sample is mixed with vitamin D antibody labeled particles forming immune complexes with total 25 OH vitamin D in the patient sample then incubated for 10 minutes at 49 C The drop of 35 uL mixture is added to the FREND Vitamin D cartridge interacting with vitamin D conjugated fluorescent nanoparticles The mixture moves via Capillary action to the detection region where fluorescent nanoparticle complexes are grabbed The fluorescence intensities from the complexes are measured and total 25 OH vitamin D concentration is calculated by the FREND System Material provided One reagent kit box contains Catalogue number 20 FREND Vitamin D cartridge s FRVD 020 20 FREND Vitamin D pretreatment tube s 20 FREND Vitamin D dilution tube s 30 Disposable pipette tip s 01 FREND Vitamin D Code chip 01 FREND
11. omatically within the cartridge once the sample has been manually loaded to the sample inlet in the cartridge and the cartridge placed into the FREND System The rate of fluorescence produced by the reaction is read at various intervals during the analysis process blank reading are subtracted after which the net rate is automatically converted to Vitamin D concentration in ng mL based upon information stored on the FREND Vitamin D Code chip This result is then output on the screen and to the optional printer It is also stored in memory on the FREND System Screen displayed for various concentration scenarios Displayed result Description Vitamin D concentration Less than 10 00 ng mL Vitamin D concentration Not less than 10 00 ng mL and not higher than 110 00 ng mL Vitamin D concentration Higher than 110 00 ng mL X 100ng dL 1ng mL 10 Limitations of the procedure 1 When used for diagnostic purposes the results obtained from this assay should be used in conjunction with other data e g symptoms results of other tests clinical impressions medical history therapy etc 2 The FREND System paired with a FREND Vitamin D cartridge is programmed to report 110 00 ng mL as the highest concentration of Vitamin D measurable without dilution The lowest measurable concentration is 10 00 ng mL the assay limit of detection 3 Specimens from patients with heterophilic antibodies such as anti mous
12. rature 18 25 C and mix gently but thoroughly before test For optimal results avoid grossly hemolytic lipidic or turbid specimens Specimens should be free of aggregated fibrin red blood cells or other particulate matter When pipetting into the FREND Vitamin D cartridge sample inlet ensure that bubbles in the sample are avoided Bubbles may restrict flow and result in an incomplete or erroneous test result Procedure Calibration There is no need for calibration to be performed by the end user as is generally required on other automated laboratory equipment All calibration statistics and information have been electronically stored on the FREND Vitamin D Code chip included in each box of FREND Vitamin D cartridges The FREND Vitamin D Code chip is specific for each manufactured lot of FREND Vitamin D cartridges Calibration information should always be checked by running external quality control samples to verify that the results obtained for Vitamin D on the FREND System using the FREND Vitamin D cartridges of a specific lot meet the laboratory criterion for acceptability Code chip installation Please refer to the FREND System user manual for more detailed instructions relative to the Code chip installation Abbreviated instructions follow here 1 Insert the FREND System electrical cord into an appropriate outlet 2 Insert the Code chip into the Code chip slot at the rear of the FREND System followin
13. rference FREND Vitamin D demonstrates lt 15 interference with the following substances at the concentrations indicated below Bilirubin Biotin Triglycerides Human albumin Hemoglobin CLS document EP7 A2 was used to analyze and calculate percent interference using serum samples at three levels of Vitamin D within the normal range Substance Concentration Bilirubin 2 5 mg dL Biotin 30 0 ng mL Triglyceride 370 0 mg dL Human albumin 10 7 g dL Hemoglobin 200 0 g dL References 1 Holick MF High prevalence of vitamin D inadequacy and implications for health Mayo Clinic Proceedings 2006 81 3 353 373 2 Bolland MJ et al The effect of vitamin D supplementation on skeletal vascular and cancer outcomes a trial sequential meta analysis The Lancet Diabetes amp Endocrinology 2014 2 4 307 320 14 3 Wolf G The discovery of vitamin D the contribution of Adolf Windaus Journal of Nutrition 2004 134 6 1299 1302 4 Holick MF Vitamin D importance in the prevention of cancers type 1 diabetes heart disease and osteoporosis The American Journal of Clinical Nutrition 2004 79 3 362 371 5 Calvo MS et al Vitamin D intake a global perspective of current status Journal of Nutrition 2005 135 2 310 316 6 Ross AC et al Dietary Reference Intakes for Calcium and Vitamin D Washington DC National Academies Press 2011 p 435 7 Svasti J et al Molecular basis for the three major forms
14. sult of test 10 When the reaction in the cartridge is completed the FREND System will automatically begin the reading process 11 When the measurements are completed the cartridge will automatically be expelled and the results displayed A Caution Do not disconnect power cord or shut off power from the FREND System while a cartridge is in the reading chamber This may cause a system error 12 If the FREND System is connected to the optional printer press the Print button and the results will be output on the printer paper 13 For more detailed instructions please refer to the FREND System User Manual Procedural notes If a specimen Vitamin D concentration is found to be greater than the linearity limit of the assay of 110 00 ng mL and a definitive result is required the specimen should be diluted with low concentration sample that has been previously measured on the FREND Vitamin D and then re assayed according to the Assay Procedure The recommended dilution for samples with an initial result of gt 110 00 ng mL is 1 2 Dilutions must be made manually and the final result on the diluted sample calculated manually by multiplying the result obtained on the diluted sample by the dilution factor X Concentration high conc sample Concentration 1 2 dilution sample X 2 Concentration low conc sample Calculation of results The FREND System performs all sample and reagent handling operations aut
15. tilized be sure these are removed from the freezer thawed naturally and then mixed gently but thoroughly prior to testing Testing should not begin on these previously frozen samples until they have reached room temperature There are no other reagents or sample preparations necessary Assay procedure 1 Prepare the FREND Vitamin D cartridge pretreatment tube dilution tube and specimen Open the pouch and place the FREND Vitamin D cartridge on the heating block 2 Transfer 35 uL of specimen to the dilution tube and mix well 3 Transfer 70 uL of diluted sample to the pretreatment tube and mix well Caution Pellet in the tube bottom should be completely dissolved 4 Put the tube in the hole of heating block and incubate for 30 minutes 5 Pipette 35 uL of the incubated sample into the sample inlet on the cartridge using a suitable micro pipette equipped with a fresh pipette tip 6 Press the Test button on the Main screen of the FREND System 7 The system moves to the Patient ID screen automatically 8 Type the Patient ID and press the Enter button to begin the test 9 Insert the cartridge into the cartridge slot using the cartridge arrow as a guide A Caution Please check the direction of the cartridge before insertion and assure the insertion is complete It is recommended to insert the cartridge after the sample injection after 30 seconds elapsed in less than 5 minutes to obtain the optimal re
16. tory Standards Institute CLSI document EP5 A2 was utilized as a guidance for precision studies on FREND Vitamin D Five human serum based panels were assayed in two replicates twice per day over a period of 20 days sample Expected Repeatability Between run Between day Within laboratory ID Value ng mL sD CV SD CV sD CV SD CV 18 00 1 443 8 0 0 348 1 9 0 282 1 6 1 511 8 4 30 00 1 643 55 0 655 2 2 0 166 0 6 1 777 5 9 60 00 3 112 5 1 0 893 1 5 1 142 1 9 3 433 5 7 90 00 4 853 5 4 2 201 25 0 787 0 9 5 386 6 0 120 00 5 486 4 6 2 606 2 2 1 333 11 6 218 5 2 almloloale Dilution linearity The range of linearity in serum was established using CLSI document EP6 A FREND Vitamin D was found to have linearity within the reportable range of 8 00 ng mL 130 00 ng mL with a mean recovery of 100 10 The dilution linearity study was performed by diluting a high concentration Vitamin D specimen with a low concentration Vitamin D specimen Results are summarized in the graphs below but may vary in individual laboratories 140 00 y 09784x 0 7883 12000 R 09994 10000 8000 6000 4000 FREND Vitamin D ng mL 2000 0 00 2000 4000 6000 8000 10000 12000 14000 Expected value ng mL Comparative analysis FREND Vitamin D was compared to the predicate device using guidelines outlined in CLSI document EP09 A2 IR Samples n 120 were measured in d
17. tridges Materials Catalogue number Refrigerator temperature storage 2 8 C FREND Vitamin D cartridges FRVD 020 FREND Vitamin D pretreatment tubes None FREND Vitamin D dilution tubes None Room temperature storage 18 25 C Pipette tips None Specimen collection and handling Human serum and plasma Lithium heparin EDTA and Citrate samples are suitable for use with FREND Vitamin D cartridges Follow instructions detailed in this package insert as well as the specimen collection tube manufacturer s instructions for specimen collection and preparation including manufacturer s instructions for centrifugation time and speed For serum a blood sample is collected aseptically without additives by venous puncture After allowing the sample to clot for 30 minutes at room temperature the collection tube should be centrifuged for 10 minutes at 3 000 rpm For plasma Lithium heparin EDTA and Citrate a venous blood sample is collected aseptically with the designated additive After allowing the specimen to sufficiently mix with anticoagulant at room temperature the sample tube can be centrifuged for 10 minutes at 3 000 rpm Samples may be stored at 2 8 for up to 6 hours prior to analysis If the analysis is scheduled to be done at some later time the sample should be stored frozen at 20 or below for future use Repeated freeze thaw cycles should be avoided Prior to assay slowly bring frozen samples to room tempe
18. uplicate on both systems Linear regression analysis was utilized to demonstrate a correlation coefficient R2 of 20 92 FREND Vitamin D ng mL 10000 8000 6000 2000 y 0978x 21536 R 09294 000 2000 4000 60 00 80 00 10000 Predicate devices ng mL Specificity FREND Testosterone demonstrates cross reactivity wit h the following related substances at the concentrations indicated bellows CLSI document EP7 A2 was used to analyze and calculate cross reactivity using serum samples at three levels of Testosterone within the normal range Substance Vitamin D concentration level Concentration Low Vitamin D2 Vitamin D3 1 25 OH 2 Vitamin D2 1 25 OH 2 Vitamin D3 3 epi 25 OH Vitamin D3 25 OH Vitamin D2 25 OH Vitamin D3 500 ng mL 500 ng mL 100 ng mL 100 ng mL 400 ng mL 25 ng mL 25 ng mL 0 0 0 2 07 0 0 0 8 103 2 Median High Cro s reactivity 0 8 08 0 8 0 5 1 0 2 2 0 9 03 1 2 1 8 106 5 96 7 99 4 97 7 113 5 Analytical sensitivity The limit of blank LoB and limit of detection LoD was determined using guidelines found in CLS document EP17 A LoB was determined from 60 replicate measurements using a calibrator A vitamin D depleted serum LoD was determined using 12 replicates measurements of five low level patient samples LoB ng mL LoD ng mL LoQ ng mL 3 84 7 07 7 07 Inte
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