This package insert follows the Pharmaceuticals, Medical devices and Other Therapeutic Products Act of Japan.
In Vitro Diagnostics
Marketing Notification No. 13A2X00197218105**Revised: January 2020 (8th edition)*Revised: July 2019 (7th edition)

This package insert must be read carefully prior to use.
hydroxyphenyl-5-phenylhydantoin, HPPH).3) Then
90% of HPPH is excreted in the urine as a glucuronic
acid conjugate of parahydroxyphenyl hydantoin, while
approximately 5% is excreted without conjugation.
Because the effective concentration range is narrow
and there is considerable individual variation of drug
absorption, metabolism, and excretion, monitoring theblood level of phenytoin is essential for adjusting thedosage during treatment.1),4),5) Toxicity of phenytoin is
dose-dependent and mainly affects the central nervoussystem. The chief symptoms of toxicity include
nystagmus, ataxia, sleepiness, diplopia, and visual
disturbance. Seizures may also be caused. As
symptoms of toxicity not involving the central nervous
system, gingival hyperplasia, anemia, andosteomalacia may occur during long-term phenytoin
therapy. As symptoms of toxicity that are notnecessarily dose-dependent, rash, adenopathy,
hepatitis, blood dyscrasias, and systemic lupus
erythematosus have also been reported.
Monitoring the blood level of phenytoin is useful for
adjusting the dosage so that the optimum clinicalresponse can be obtained without symptoms of toxicity.

Phenytoin assay kit(Classification No.: 30402000)

Nanopia TDM Phenytoin
General Precautions
1. This product is for in vitro diagnostic use, and mustnot be used for any other purposes.
2. Clinicians should make a comprehensive clinicaldecision based on assay results in conjunction withclinical symptoms and other examination results.
3. This product should be used only as directed in thispackage insert. Reliability of results cannot be
guaranteed if there are any deviations from theinstructions in this package insert.
4. If the reagent accidentally comes in contact witheyes and/or mouth, rinse immediately with ample
water as first aid, and consult the doctor if required.
5. Carefully read the operating instructions for eachtype of automated analyzers prior to using thisproduct. Parameters for each type of analyzers areavailable, and can be requested from SEKISUIMEDICAL CO., LTD. if required.
6. Perform a quality control test prior to assay toensure accuracy.
7. PHT Antibody Solution 1 contains human-derived
components that have been shown to be negative for
HBs antigens, HIV antibodies (AIDS virusantibodies), and HCV antibodies. However, thesereagents (as well as the samples) should beconsidered potentially infectious and handled withgreat care.

Assay Principle
1. Assay Principle
When a certain amount of anti-phenytoin antibody
is added and reacted with a sample, consumptionof the antibody depends on its content in thesample. When phenytoin-coated latex is added,
residual anti-phenytoin antibody reacts with thelatex and forms aggregates.
Since the extent of aggregation depends on thephenytoin concentration in the sample, thephenytoin concentration can be determined bymeasuring aggregation as the change ofabsorbance.

Description (Kit Components)
Component: IngredientsPHT Antibody Solution 1:
Anti-phenytoin mousemonoclonal antibody
PHT Latex Reagent 2:Phenytoin-coated latex

Sample (phenytoin) + Anti-phenytoin antibody──────▶ Antigen-antibody reaction
Unreacted anti-phenytoin antibody + Phenytoincoated latex───▶ Aggregation by antigen-antibody reaction
2. Features
1) Because a highly specific monoclonal
antibody is used, this product shows excellent
sensitivity and accuracy.
2) Liquid reagents, ready-to-use.
3) Applicable to various automated analyzers.

Intended Use
Measurement of phenytoin in serum or plasma
Phenytoin (diphenylhydantoin) is one of the drugs
used to treat epilepsy. It is most widely prescribed as
an anticonvulsant for grand mal seizures (mostlykinetic), focal seizures, and temporal lobe epilepsy.1)
Phenytoin is usually administered orally. It is almost
completely absorbed (85−95%) from the small
intestine.2) Approximately 87−94% of phenytoin in thebloodstream is bound to plasma protein and ispharmacologically inactive, while unbound (free)phenytoin has pharmacological activity. If phenytoin
is administered in combination with various otherdrugs or is administered to patients with complications,the free phenytoin level may increase to induce
toxicity. From 70−80% of a dose of phenytoin
undergoes hydroxylation in the liver and ismetabolized to an inactive metabolite (5-＊＊Procedural Precautions ＊1. Properties of Samples and Sampling Methods
1) Samples
Serum and plasma (heparin plasma, EDTA
plasma and citrated plasma) may be used.
2) Storage of samples
If serum or plasma samples cannot bemeasured on the day of separation, store themas follows. Avoid repetition of freezing and1

thawing.
2–8C: for tests within 7 days≤ -20C: for tests within 4 months
Bring samples to room temperature (15–30C)
before use.
3) Caution must be exercised, because aseparating agent, etc. in the blood collection
tube may affect assay values.6)4) Sampling should be performed after
removing insoluble matter from the sample.
2. Interfering substances
1) Assay results are not affected by free bilirubin(up to 20 mg/dL), conjugated bilirubin (up to 20mg/dL), hemoglobin (up to 500 mg/dL),ascorbic acid (up to 50 mg/dL), formazinturbidity (up to 2500 FTU), or rheumatoid
factors (up to 200IU/mL).
2) Because mouse antibody is used in the assay,
artifactual elevation of results may occur if thesample contains human anti-mouse antibody. In
this case, perform re-measurement by anothermethod.
3) Cross-reactivity
The following table summarizes drugs related
to phenytoin and cross-reactivity between
phenytoin and other drugs.

Substance

Concentration
tested(µg/mL)
500

for calibration.
2) Precautions for assay range
If the concentration of a target substance in thesample exceeds the measurement range, dilutethe sample with a separately sold diluent(manufactured by SEKISUI MEDICAL CO.,LTD.), and perform re-measurement.
Dosage/Administration (Assay Procedure)
1. Preparation of reagents
Reagent (1): PHT Antibody Solution 1 is readyto use.
Reagent (2): PHT Latex Reagent 2 is ready touse.
Before using this product, gently invert the PHT
Latex Reagent 2 bottle to mix it thoroughly, andcheck that there are no bubbles.
2. Assay Procedure
37C37CReagent
Sample Reagent
Measurement+(1)
(2)
2 μL
270
70 (Absorbance I※)
270 μL100 μLsecsec37CMeasurement247 (Absorbance II※)sec
Calculation ofconcentration

Crossreactivity (%)※

Absorbance I and II: Absorbance at 700 nm

Calibration material: TDM Calibrator for Nanopia(manufacture's assigned value)

5-(p-Methylphenyl)-55.60
phenylhydantoin
Amitriptyline
3000
0.01
Amobarbital
100
0.05
Carbamazepine
500
0.08
Carbamazepine-10,111000
0.06
epoxideClorazepate
2000
0.03
Chlordiazepoxide
2000
0.01
Chlorpromazine
2500
0.00
Diazepam
2000
0.02
Ethosuximide
1000
0.02
Ethotoin1000
0.17
Glutethimide
1000
0.00
5-(p-Hydroxyphenyl)-5500
2.46
phenylhydantoin (HPPH)
Hydantoin
2000
0.02
Imipramine
4000
0.01
Mephenytoin
3000
0.04
Mephobarbital
1000
0.07
Methsucimide
5000
0.01
Oxaprozin
500
0.10
2-Phenyl-21000
0.01
ethylmalonamide
Pentobarbital
1000
0.03
Phenobarbital
2000
0.00
Phensuximide
2000
0.04
p-Hydroxyphenobarbital
2000
0.00
Primidone
1000
0.04
Promethazine
1500
0.01
Secobarbital
2000
0.02
Valproic acid
7000
0.00
3. Others
1) Always use TDM Calibrator for Nanopia

Assessment of Assay Results ＊＊1. Reference standard range
The therapeutic concentration of phenytoin has
been reported to be 10−20 μg/mL in adults andchildren, 8−15 μg/mL in newborns.7) Assay results
should be assessed while taking into consideration
the patient’s clinical findings and other laboratorydata.
It is recommended that the blood concentration ofphenytoin should be measured in a sufficientnumber of samples for statistical analysis and thatits clinical effective concentration should bedetermined by each medical institution.
2. There may be reactions or interfering reactions
with non-target substances. If assay results appear
to be unreliable, repeat the measurement (ifnecessary, after dilution) or try another analyticalmethods.
Performance ＊＊1. Sensitivity
1) The change of absorbance of the standardsolution (0 μg/mL) per minute is 0.05−0.21.
2) The ratio between the change of absorbance perminute with the standard solution (0 µg/mL)
and that with another standard solution (2.5µg/mL) is 60−95%.
2. Accuracy: 80–120 % of the expected assay value
3. Within-run Reproducibility:
Coefficient of variation ≤ 10 %(Test methods used for 1.–3. are in-house methods.)
4. Measurement Range8): (On Hitachi 7170S2

automated analyzer)
0.7–40 μg/mL
5. Correlation8)
1) Serum N=65
r=0.997
y=1.03x+0.06
Control method: Approved in vitro diagnostic(enzyme immunoassay)
2) Plasma
N=98
r=0.995 y=0.95x+0.16
Control method: Approved in vitro diagnostic(enzyme immunoassay)
6. Standard Material
Phenytoin (U.S. Pharmacopoeia)

2. Shelf life: 19 months from the date of manufacture(The expiration date is printed on the outer
package.)
Packaging
NamePackage
PHT Antibody
1 × 54 mL
Solution 1
Nanopia TDM
Phenytoin
PHT Latex
1 × 20 mL
Reagent 2Constituent reagents are available in otherconfigurations. For further details please contact
SEKISUI MEDICAL CO., LTD.

Precautions for Use or Handling
1. Precautions for Handling (to Ensure Safety)
1) All samples used in the test should be handled asa material possibly infected with HIV, HBV,HCV, or other viruses. To prevent infection, use
disposable gloves and avoid mouth pipetting
during the test.
2) Sodium azide is added as an antiseptic agent in
the PHT Antibody Solution 1 and PHT Latex
Reagent 2. Therefore, if the reagent comes inaccidentally contact with eyes, mouth or skin,rinse immediately with ample water as first aid,and consult the doctor if required.
2. Precautions for use
1) This product should be stored as directed,without freezing. Freezing can deteriorate thereagents, which can produce inaccurate results.
Therefore, avoid using the reagents which havebeen previously frozen.
2) Do not use expired reagents. Use of suchreagents cannot guarantee the reliability ofmeasurement values.
3) Do not replenish the reagents.
4) Do not mix materials from different kit lotnumbers.
5) Do not perform the assay under direct sunlight
3. Precautions for Disposal
1) Before disposal, used samples and theircontainers must be immersed in sodium
hypochlorite solution at a concentration ofgreater than 0.1% for longer than 1 hour or
autoclaved at 121ºC for 20 minutes.
2) To prevent infections from spilled samples orsolutions containing samples, wipe the spilledarea thoroughly with disinfectants such as
sodium hypochlorite solution at a concentrationof greater than 0.1%.
3) The reagents and treated samples should bediscarded as medical waste or industrial waste
according to the waste disposal regulations.
4) The reagents should be disposed of inaccordance with the Water Pollution Control act
or related regulations.
5) Sodium azide has been added as an antiseptic
agent in the PHT Antibody Solution 1 and PHT
Latex Reagent 2. It can react with lead or copper
pipes to produce the highly explosive metal azide.
Therefore, the reagent should be flushed withlarge amounts of water during disposal.
4. Other precautions
Do not use the containers for other purposes.

References ＊＊1) Buchthal F., Lennox-Buchthal M. A. ：Diphenylhydantoin ： Relation of anticonvulsant
effect to concentration in serum, in Woodbury, D.
M., Penry J. K., Schmidt K. P. （ eds. ） :Antiepileptic Drugs, Raven Press, New York, pp.
193−209, 1972.
2) Finn A. L., Olanow C. W.：Phenytoin：Therapeutic
use and serum concentration monitoring, in Taylor,W. J., Fin, A. L. (eds.): Individualizing Drug
Therapy ：Practical Application of Drug
Monitoring, Vol 2, Gross, Townsend, Frank, Inc.,New York, pp. 63−85, 1981.
3) Jusko W. J.：Bioavailability and disposition kineticsof phenytoin in man, in Kellaway, P., Petersen, I.
（eds.): Quantitative Analytic Studies in Epilepsy,Raven Press, New York, pp. 115−136, 1976.
4) Kutt H. ： Diphenylhydantoin ： Relation of plasma
levels to clinical control, in Woodbury, D. M.,
Penry, J. K., Schmidt, K. P. （eds.）: AntiepilepticDrugs. Raven Press, New York, pp. 211−218, 1981.
5) Buchthal F., Svensmark O.：Serum concentrationsof diphenylhydantoin （ Phenytoin ） andphenobarbital and their relation to therapeutic andtoxic effects, Psychiat. Neurol. Neurochir., 74:117−136, 1971.
6) Sawada T. et al.: J Med Pharm Sci, 51(1), 131−141,2004.
7) The Japanese Society of Therapeutic Drug
Monitoring. [Guideline for Therapeutic Drug
Monitoring (TDM) for Antiepileptic Drug 2018].
Tokyo: Kanehara Shuppan; 2018. pp. 8-9, pp. 3239.
8) In house data, SEKISUI MEDICAL CO., LTD.
Contact
SEKISUI MEDICAL CO., LTD.
international@sekisui.com

Manufacturer
SEKISUI MEDICAL CO., LTD.
1-3, Nihonbashi 2-chome, Chuo-ku, Tokyo, Japan

Storage and Shelf Life
1. Storage temperature: 2–8C

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