Synthesis and preclinical pharmacological evaluation of a novel 99mTc–shikonin as a potential tumor imaging agent

Shikonin was isolated from Ratanjot pigment then the obtained shikonin was well characterized. This study is aimed to optimize radiolabeling yield of shikonin with ^99mTc with respect to factors that affect the reaction conditions such as shikonin amount, SnCl₂·2H₂O amount, reaction time and pH of the reaction mixture. In vitro stability of the radiolabeled complex was checked and it was found to be stable for up to 6?h. Biodistribution studies showed that, ^99mTc?Cshikonin accumulate in tumor sites with higher T/NT than other currently available ^99mTc(CO)₃-VIP, ^99mTc?Cnitroimidazole analogues and ^99mTc?Cpolyamine analogues indicating that shikonin deliver ^99mTc to the tumor sites with a percentage sufficient for imaging and can overcome many drawbacks of other radiopharmaceuticals used for tumor imaging.     

Evaluation and comparison of 99mTc-labeled d-glucosamine derivatives with different 99mTc cores as potential tumor imaging agents

Isocyanide is a strong coordination ligand that can coordinate with [^99mTc(I)(CO)₃]⁺ core and [^99mTc(I)]⁺ core to produce stable ^99mTc complexes, therefore developing a ^99mTc-labeled isocyanide complex for single-photon emission computed tomography (SPECT) imaging is considered to be of great interest. In order to develop potential tumor imaging agents with satisfied tumor uptake and suitable pharmacokinetic properties in vivo, a novel d -glucosamine isocyanide derivative, 4-isocyano-N-(2,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-3-yl)butanamide (CN3DG), was synthesized and radiolabeled with [^99mTc(I)]⁺ and [^99mTc(CO)₃]⁺ cores to obtain [^99mTc(CN3DG)₆]⁺ and [^99mTc(CO)₃(CN3DG)₃]⁺ in high radiolabeling yields (>95%). Both of the complexes showed good hydrophilicity and great stability in vitro. Cell uptake studies performed in S180 cells demonstrated they were transported into cells by glucose transporters. Biodistribution studies of the two complexes in mice bearing S180 tumor showed they had high tumor uptakes and rapid clearance from muscle and blood so that the tumor/blood and tumor/muscle ratios were high. By comparison, [^99mTc(CN3DG)₆]⁺ was superior to [^99mTc(CO)₃(CN3DG)₃]⁺ in regard to tumor uptake, tumor/blood and tumor/liver ratios. S180 tumors could be seen clearly from the SPECT/CT images with [^99mTc(CN3DG)₆]⁺. Considering its favorable properties, [^99mTc(CN3DG)₆]⁺ would be a promising tumor imaging agent and needs to be further studied.     

Characterization of 99mTc(CO)3-iminodiacetic acid (IDA) and its comparison with 99mTc-(IDA)2 using compounds for hepatobiliary scintigraphy

The organometallic precursor of fac-[^99mTc(CO)₃(H₂O)₃]⁺ has attracted much attention because of the robustness and small size of Tc(I)-tricarbonyl complexes compared to Tc(V) complexes and the good labeling affinity with a variety of donor atoms. Among various ligand systems, an iminodiacetic acid (IDA) was proven as a good chelating group to form a Tc(III)-compelx as well as has been shown its potential as a chelating system for fac-[^99mTc(CO)₃] precursor. In an attempt to confirm the similarity and the difference between ^99mTc(CO)₃-IDA and ^99mTc-(IDA)₂-complex, M(CO)₃-IDA (M = ^99mTc, Re) complexes of disofenin, mebrofenin and N-(3-iodo-2,4,6-trimethyl phenylcarbamoylmethyl) iminodiacetic acid were prepared, and the biological evaluation of ^99mTc(CO)₃-disofenin was performed. The ^99mTc(CO)₃-IDA complexes were prepared with a high radiolabeling yield (>98%) in a quantitative manner and showed a negative charge. The in vivo pharmacokinetic behavior of ^99mTc(CO)₃-disofenin showed a similar biological activity to ^99mTc-(disofenin)₂ in that those complexes were quickly cleared from the blood by the hepatocytes and excreted into the gallbladder and intestine. Accordingly, the ^99mTc(CO)₃-IDA derivatives of disofenin and mebrofenin might be used as hepatobiliary imaging agents. Since an IDA is a promising chelator for ^99mTc-based radiopharmaceutical and the biological properties of ^99mTc(CO)₃-IDA derivative shows similar to that of ^99mTc-complex, a biomolecule containing IDA can be freely radiolabeled with fac-[^99mTc(CO)₃]-precursor or ^99mTc. However, the radiolabeling efficiency and the biological behavior demonstrates the favorable properties of ^99mTc(CO)₃-IDA compound for the development of a new imaging agent.     

Synthesis and biological distribution of 99mTc–norfloxacin complex, a novel agent for detecting sites of infection

The optimization of the radiolabeling yield of ciprofloxacin analogous, norfloxacin, with technetium-99m (^99mTc) was described. Dependence of the labeling yield of ^99mTc–norfloxacin complex on the concentration of norfloxacin, SnCl₂·2H₂O content, pH of the reaction mixture and reaction time was studied. Norfloxacin was labeled with ^99mTc at pH 3 with a labeling yield of 95.4% by using 5 mg norfloxacin, 50 μg SnCl₂·2H₂O and 30 min reaction time. The formed ^99mTc–norfloxacin complex was stable for a time up to 3 h. Biological distribution of ^99mTc–norfloxacin complex was investigated in experimentally induced inflammation rats using Staphylococcus aureus (bacterial infection model) and heat killed Staphylococcus aureus and turpentine oil (sterile inflammation model). In case of bacterial infection, the T/NT value for ^99mTc–norfloxacin complex was found to be 6.9 ± 0.4 which was higher than that of the commercially available ^99mTc–ciprofloxacin under the same experimental condition.     

Radiosynthesis and evaluation of novel [99mTc(I)]+ and [99mTc(I)(CO)3]+ complexes with a 4-nitroimidazole isocyanide for imaging tumor hypoxia

[^99mTc(I)]⁺ and [^99mTc(I)(CO)₃]⁺ complexes with isocyanide exhibit high stability, which makes them suitable platforms to develop novel ^99mTc radiopharmaceuticals. To develop novel ^99mTc radiotracers for imaging hypoxia, in this study, a novel L ligand (4-nitroimidazole isocyanide derivative) was synthesized and labelled using [^99mTc(I)]⁺ core and [^99mTc(I)(CO)₃]⁺ core to produce [^99mTc(L)₆]⁺ and [^99mTc(CO)₃(L)₃]⁺ with high yields. To verify the structure of the ^99mTc complexes, corresponding rhenium analogues were synthesized and characterized. Both of the ^99mTc complexes were stable and hydrophilic. in vitro cellular uptake results showed they could exhibit good hypoxic selectivity. The evaluation of biodistribution in mice bearing S180 tumors indicated both of them could accumulate in tumor. Between them, [^99mTc(L)₆]⁺ exhibited higher tumor uptake and tumor/non-target ratio than [^99mTc(CO)₃(L)₃]⁺. Further, single photon emission computed tomography (SPECT) imaging studies of [^99mTc(L)₆]⁺ indicated an obvious accumulation in tumor and the value of the region-of-interest (ROI) ratio of the uptake for the tumor site to the corresponding non-tumor region was 5.64 ± 0.52. The above results suggested [^99mTc(L)₆]⁺ would be a potential tracer for imaging tumor hypoxia.     

Practicality of Tetragonal Nano-Zirconia as a Prospective Sorbent in the Preparation of 99Mo/99mTc Generator for Biomedical Applications

The feasibility of using tetragonal nano-zirconia (t-ZrO₂) as an effective sorbent for developing a ⁹⁹Mo/^99mTc chromatographic generator was demonstrated. The structural characteristics of the sorbent matrix were investigated by different analytical techniques such as XRD, BET surface area analysis, FT-IR, TEM etc. The material synthesized was nanocrystalline, in tetragonal phase with an average particle size of ~7 nm and a large surface area of 340 m² g^?1. The equilibrium sorption capacity of t-ZrO₂ is >250 mg Mo g^?1. The present study indicates that ⁹⁹Mo is both strongly and selectively retained by t-ZrO₂ at acidic pH and ^99mTc could be readily eluted from it, using 0.9% NaCl solution. A 9.25 GBq (250 mCi) t-ZrO₂ based chromatographic ⁹⁹Mo/^99mTc generator was developed and its performance was repeatedly evaluated for 10 days. ^99mTc could be eluted with >85% yield having acceptable radionuclidic, radiochemical and chemical purity for clinical applications. The compatibility of the product in the preparation of ^99mTc labeled formulations such as ^99mTc-EC and ^99mTc-DMSA was evaluated and found to be satisfactory.     

Effect of chemical condition of technetium-99m sodium pertechnetate on the active kit components and radiolabeling yield of Tru-Scint® ADTM monoclonal antibody kit

The chemical condition of^99mTc eluate obtained from a⁹⁹Mo-^99mTc generator is a function of the source, time elapsed after elution and age of the eluate. The radiochemical purity and stability of^99mTc labeled MAb-170 (Tru-Scint^®AD^TM, photoactivated monoclonal antibody kit) preparations was evaluated comparing pertechnetate source of known age and elution history. The effect of H₂O₂, a radiolytic impurity in^99mTc eluates, on the active kit components stannous ion and photoactivated MAb and radiolabeling, yield has been investigated. The lyophilized Tru-Scint^® AD^TM kit has been labeled with 20 to 80 mCi in 0.5 to 4.0 ml of Sodium Pertechnetate^99mTc Injection, USP. The eluates were obtained from three brands of generators and used up to six hours after elution. The kits were reconstituted either with Sodium Pertechnetate^99mTc Injection, USP or Sodium Chloride Injection, USP, 0.9% containing known amounts of H₂O₂. The reconstituted kits were analyzed for radiolabeling yield and radiochemical impurities, stannous ion and protein sulfhydryl group. The results indicated that the radiolabeling yield is a function of both the chemical condition of^99mTc eluate, generator brand and the radiolabeling parameters like reconstitution volume and activity. The observed radiolabeling yield differences did not depend on the amount of chemical technetium in the eluate. The major radiochemical impurities at 15-minute post labeling have been identified as the^99mTc-buffer complex and column adsorbed reduced^99mTc (^99mTc-Ad) species and not the unreduced^99mTcO ₄ ^– .     

Preparation and biological evaluation of Tc-CO-MIBI as myocardial perfusion imaging agent

^99mTc-Sestamibi has been playing an important role in the cardiac imaging for the last decades. Previously, we reported that [^99mTc(CO)₃(MIBI)₃]⁺ demonstrated a significant location in myocardium with a lower liver uptake as compared with ^99mTc-Sestamibi. In this work, we found that new [^99mTc(CO)₂(MIBI)₄]⁺ could be prepared with high radiochemical purity. The inter-transformations between [^99mTc(CO)₃(H₂O)(MIBI)₂]⁺, [^99mTc(CO)₃(MIBI)₃]⁺, and [^99mTc(CO)₂(MIBI)₄]⁺ were investigated and biodistribution was performed to evaluate the [^99mTc(CO)₂(MIBI)₄]⁺ as a myocardial perfusion imaging agent. The results showed that one more CO was replaced by MIBI slowing down the pharmacokinetics. The structure characterization was performed on their corresponding rhenium complexes, and the results indicated that there were differences between ^99mTc-CO-MIBI and Re-CO-MIBI in preparation and hydrophobic characteristics.     

Synthesis and preliminary biological evaluation of 99mTc(CO)3-labeled pegylated 2-nitroimidazoles

This work reports the synthesis, radiolabeling and preliminary biodistribution results in tumor-bearing mice of ^99mTc(CO)₃-labeled pegylated (PEG) 2-nitroimidazoles for tumor hypoxia imaging. The novel 2-nitroimidazole derivatives were successfully synthesized by conjugation of tridendate chelators to 2-nitroimidazole via PEG₃ linker. Radiolabeling was performed in high yield with [^99mTc(CO)₃]⁺ core to get cationic [^99mTc(CO)₃(BPA-PEG₃-NIM)]⁺, neutral [^99mTc(CO)₃(AOPA-PEG₃-NIM)] and anionic [^99mTc(CO)₃(IDA-PEG₃-NIM)]^? respectively, all of which were hydrophilic and stable at room temperature. Biodistribution studies in tumor-bearing mice showed that ^99mTc(CO)₃-labeled pegylated 2-nitroimidazoles accumulated in the tumor with low uptake. ^99mTc-chelate and charge had significant impact on partition coefficient, radiotracer tumor uptake and pharmacokinetic properties. The results indicate the need for synthetic modification of the parent 2-nitroimidazole derivatives and the ^99mTc-chelate with a view to improve the tumor targeting efficacy and in vivo kinetic profiles.     

Rapid and sensitive determination of pertechnetate in99Mo/99mTc generator eluates by reversed-phase high-performance liquid chromatography

Reversed-phase high-performance liquid chromatography with u.v. detection was applied for rapid and sensitive determination of pertechnetate in⁹⁹Mo/^99mTc generator eluates, using a mixture solvent of acetonitrile and 0.04M aqueous acetate buffer (1/1) containing a few volume percentage of 0.5 M tetra-n-butylammonium hydroxide as the mobile phase. Employing a -bondapak C₁₃ column, the TcO ₄ ^– species was separated, monitored with absorbance at 254 nm, and observed at the retention time of 3.5 min. The detection limit was found to be 5.2·10^–10 g of Tc for each injection. Total Tc contents in the^99mTc eluates from clinically-used⁹⁹Mo/^99mTc generator were analyzed by this technique. The^99mTc (⁹⁹Tc) species was separated from the contaminant⁹⁹Mo. This method was found to be useful for the purification of^99mTc (⁹⁹Tc) as well as the determination of total Tc content.     

Determination of calibration curves for <Superscript>131</Superscript>I in thyroid tumour metabolic radiotherapy and other radionuclides used in SPECT imaging

Large columns containing aluminum oxide (Al₂O₃) or gel (e.g. zirconium molybdate) are needed to prepare ⁹⁸Mo(n,γ)⁹⁹Mo→^99mTc column chromatographic generators that results in large elution volumes containing relatively high ⁹⁹Mo impurity and low concentrations of ^99mTc. The decrease in radioactive concentration or specific volume concentration of ^99mTc places a limitation on some pharmaceutical kits (DTPA, MIBI, ECD, etc.) or clinical procedures. We report on the post elution concentration of ^99mTc using in house prepared lead cation-exchange and alumina columns. Using these columns high bolus volumes (10–60 mL 0.02M sodium sulfate) of ^99mTc can conveniently be concentrated in 1 mL of physiological saline. This approach also works very effectively to prepare high specific volume solutions of ^99mTc-pertechnetate from a fission based ⁹⁹Mo/^99mTc generator in the second week of its normal working life.     

In vitro incorporation studies of 99mTc–alendronate sodium at different bone cell lines

Bisphosphonates can be labeled with Technetium-99m (^99mTc) and are used for bone imaging because of their good localization in the skeleton and rapid clearance from soft tissues. Over the last decades bone scintigraphy has been used extensively in the evaluation of oncological patients to provide information about the sites of bone lesions, their prognosis and the effectiveness of therapy by showing the sequential changes in tracer uptake. Since the lesion visualization and lesion/bone ratio are important utilities for a bone scanning radiopharmaceutic; in this study incorporation of ^99mTc labeled alendronate sodium (^99mTc–ALD) was evaluated in U₂OS (human bone osteosarcoma) and NCI-H209 (human bone carcinoma) cell lines. ALD was directly labeled by ^99mTc, radiochemical purity and stability of the complex were analyzed by radioactive thin layer chromatography and radioactive high performance liquid chromatography studies. For cell incorporation study, NCI-H209 and U₂OS cell lines were used with standard cell culture methods. The six well plates were used for all experiments and the integrity of each cell monolayer was checked by measuring its transepithelial electrical resistance (TEER) with an epithelial voltammeter. Results confirmed that ALD was successfully radiolabeled with ^99mTc. ^99mTc–ALD incorporated with NCI-H209 and U₂OS cells. The uptake percentages of ^99mTc–ALD in NCI-H209 and U₂OS cell lines were found significantly different. Since ^99mTc–ALD highly uptake in cancer cell line, the results demonstrated that radiolabeled ALD may be a promising agent for bone cancer diagnosis.     

<Superscript>99m</Superscript>Tc(CO)<Subscript>3</Subscript>–AOPA colchicine conjugate as a potential tumor imaging agent

This work reports the synthesis, radiolabeling and preliminary biodistribution results in tumor-bearing mice of the ^99mTc(CO)₃–AOPA colchicine conjugate. The novel ligand was successfully synthesized by conjugation of N-(acetyloxy)-2-picolylamino (AOPA) to deacetylcolchicine via a short carbonyl-methylene linker. Radiolabeling was performed in high yield with [^99mTc(CO)₃]⁺ core. ^99mTc(CO)₃–AOPA colchicine conjugate was hydrophilic and was stable at room temperature. Biodistribution studies in tumor-bearing mice showed that ^99mTc(CO)₃–AOPA colchicine conjugate accumulated in the tumor with good uptake and retention. However, its clearance from normal organs was not so fast, resulting in poor T/NT ratios. Further modification on the linker or/and ^99mTc-chelate to improve the tumor targeting efficacy and in vivo kinetic profiles is currently in progress.     

The interaction of <Superscript>99m</Superscript>Tc with pyrimidine compounds

The reaction of ^99mTc of different oxidation states (+7, +4) with 2-thiouracil and 5-nitrobarbituric acid have been studied at different temperatures, pH and concentrations. The reaction mixtures have been analyzed at different times using thin layer chromatography (TLC) and a radio detector to show the peaks at the plates. ^99mTc is obtained from the Mo generators with oxidation state (+7). The use of SnCl₂ as a reducing agent gave ^99mTc with oxidation state (+4). It is very difficult to separate the complexes formed from the reactions in very small concentration. The percentage of ^99mTc and its oxidation state involved in the complexes can be determined. The labeling efficiencies (percentage of complex) in the reaction of ^99mTc⁺⁷ with 5-nitro-barbituric-acid increases mostly at pH  10. Both oxidation states of ^99mTc(+7, +4) can be detected at pH’s 4 and 10, but at pH  4, the reduced form ^99mTCO₂, is more pronounced. At pH  7 no complexes were detected and most of ^99mTc remains as ^99mTCO₄ ⁻ . By increasing the ligand concentration, the labeling efficiencies of the complex increases. For the reaction of ^99mTc of oxidation states (+4, +7) with 2-thiouracil at different temperatures and analytical times it is concluded that several complexes with different R_f values were observed in equilibrium and most of these complexes were unstable.     

Preparation,molecular modeling and biodistribution of 99mTc-phytochlorin complex

Phytochlorin [21H, 23H-Porphine-7-propanoicacid, 3-carboxy-5-(carboxymethyl)13-ethenyl-18-ethyl-7,8-dihydro-2,8,12,17-tetramethyl-,(7S,8S)] was labeled with ^99mTc and the factors affecting the labeling yield of ^99mTc-phytochlorin complex were studied in details. At pH 10, ^99mTc-phytochlorin complex was obtained with a high radiochemical yield of 98.4 ± 0.6 % by adding ^99mTc to 100 mg phytochlorin in the presence of 75 μg SnCl₂·2H₂O after 30 min reaction time. The molecular modeling study showed that the structure of ^99mTc-phytochlorin complex presents nearly linear HO–Tc–OH unit with an angle of 179.27° and a coplanar Tc(N1N2N3N4) unit. Biodistribution of ^99mTc-phytochlorin complex in tumor bearing mice showed high T/NT ratio (T/NT = 3.65 at 90 min post injection). This preclinical study showed that ^99mTc-phytochlorin complex is a potential selective radiotracer for solid tumor imaging and afford it as a new radiopharmaceutical suitable to proceed through the clinical trials for tumor imaging.     

Comparison of ‘classic’ Tc-DTPA, Tc(CO)3-DTPA and Tc(CO)2(NO)-DTPA

Diethylenetriamine pentaacetic acid (DTPA) was labeled with ^99mTc in three different ways, resulting in ‘classic’ ^99mTc-DTPA, ^99mTc(CO)₃-DTPA and ^99mTc(CO)₂(NO)-DTPA. The biodistribution of the formed DTPA-complexes was studied in mice with a special emphasis on the behavior of the novel tricarbonyl and dicarbonyl-nitrosyl complexes, which was clearly differing from that of ‘classic’ ^99mTc-DTPA. The conversion of a Tc-tricarbonyl complex to a Tc-dicarbonyl-nitrosyl complex using NO⁺ reagents offers a synthetic tool for preparing a novel class of ^99mTc labeled compounds.     

Novel 99mTc labeled σ receptor ligand as a potential tumor imaging agent

A novel ^99mTc labeled complex, [N-[2-((2-oxo-2-(4-(3-phenylpropyl)piperazin-1-yl)ethyl) (2-mercaptoethyl)amino)acetyl]-2-aminoethanethiolato]Technetium(V) oxide (PPPE-MAMA’-^99mTcO) ([ ^99m Tc]-2) has been designed and prepared based on the integrated approach. The corresponding rhenium complex (PPPE-MAMA’-ReO)(Re-2) has been prepared and characterized. In vitro competition binding assays show moderate affinity of Re-2 towards σ₁ and σ₂ receptors with K _i values of 8.67 ± 0.07 and 5.71 ± 1.88 μmol, respectively. Planar images obtained at 0.5 h, 4 h, 20 h after i.v. injection indicate the accumulation of [ ^99m Tc]-2 in MCF-7 human breast tumor bearing mice at 20 h. Furthermore, the accumulation of [ ^99m Tc]-2 has been inhibited at 20 h after co-injection of [ ^99m Tc]-2 plus haloperidol (1 mg/kg). Biodistribution studies of [ ^99m Tc]-2 display an in vivo tumor uptake of 0.14% ± 0.01% ID/g at 24 h post i.v. injection with a tumor/muscle ratio of 6.02 ± 0.87. The above results suggest that [ ^99m Tc]-2, derived from a previously published lead compound, retains certain tumor uptake and affinity for σ receptors. [ ^99m Tc]-2 may be used as a basis for further structural modifications to develop tumor imaging agents with high affinity for σ receptors.     

Neutron resonance capture analysis and applications

This paper addresses the development of two new radiopharmaceuticals for infection imaging. The optimization of the labeling yield of ciprofloxacin analogous, lomefloxacin and ofloxacin, with ^99mTc is described. ^99mTc-lomefloxacin was obtained with a radiochemical yield of 93.6% by adding ^99mTc to 2.5 mg lomefloxacin in the presence of 50 μg SnCl₂ while ^99mTc-ofloxacin was obtained (96.6%) by adding ^99mTc to 2 mg ofloxacin in the presence of 50 μg SnCl₂. Biodistribution studies in rats were carried out in experimentally induced infection in the left thigh using Staphylococcus aureus. Both thighs of the rats were dissected and counted and the ratio of bacterial infected thigh/contralateral thigh was then evaluated. ^99mTc-lomefloxacin showed higher uptake (T/NT = 6.5±0.5) in the infectious lesion than ^99mTc-ofloxacin (T/NT = 4.3±0.6) and abscess-to-muscle ratios for both preparations were higher than that of ^99mTc-ciprofloxacin (T/NT = 3.8±0.8), indicating that ^99mTc-lomefloxacin could be used for infection imaging.     

Effective synthesis of Tc tricarbonyl complexes by microwave heating

Technetium-99m (^99mTc) is one of the most frequently used nuclides for single-photon emission computed tomography (SPECT) imaging because of its radiochemical characteristics, such as gamma emission of suitable energy (141 keV) and adequate half-life (6.01 h). Although triaquatricarbonyl ^99mTc cation ([^99mTc(CO)₃(H₂O)₃]⁺) has several advantages as a ^99mTc-labeling agent, e.g., compact chelate size, chelate stability, and simplicity of preparation, its synthetic protocols should be improved. Because microwave heating is a convenient method for synthetic reactions, we studied the effect of microwave irradiation on the synthesis of ^99mTc tricarbonyl complexes. We found several factors beneficial for the preparation of nuclear medicines. In particular, microwave heating promoted one-pot syntheses of ^99mTc tricarbonyl chelates in a short time. In addition, the ^99mTc tricarbonyl complex could be obtained using low concentrations of ligands.     

Dose rate determination of gamma rays emitted by Thallium-201 and technetium-99m using a modified horseradish peroxidase based biosensing system

In the presence of low energy gamma emitter radioisotopes of thallium-201 (²⁰¹T1) or technetium-99m (^99mTc), H₂O₂ was generated via radiolysis of water. The produced H₂O₂ was amperometrically determined using an anthraquinone 2-carboxylic acid modified horseradish peroxidase on glassy carbon electrode. In the presence of each radioisotope, the cathodic current produced due to the amperometric detection of H₂O₂, was designated as biosensor response. At the applied potential of -550 mV (vs. Ag/AgCl), the biosensor showed the sensitivities of 1.937 and 2.278 nA h ?Gy^-1 towards H₂O₂ produced by ²⁰¹Tl and ^99mTc respectively. Finally, the calibration curves for dose rate determination of ²⁰¹Tl and ^99mTc have been presented and the correlations between biosensor response to H₂O₂ and the gamma emitter dose rates for ²⁰¹Tl and ^99mTc are established.