A novel method for labelling of leukocytes with technetium-99m and its comparative evaluation for abscess scintigraphy

A new method, based on the pretreatment of leukocytes with glucoheptonate prior to treating with reduced ^99mTc, has been developed for the preparation of ^99mTc labelled leukocytes. The leukocytes labelled with a ^99mTc concentration (5.59%/g tissue) similar to that of ¹¹¹In-leukocytes (6.27%/g tissue), in the experimental abscess were in rat thigh. Concentration of ^99mTc-leukocytes in blood at 24 h was only about 35% as compared to that of ¹¹¹In-leukocytes. Biodistribution in the rat organs was similar in both cases, except in the liver where ^99mTc-leukocytes exhibited about 4-fold greater concentration. Images of experimental abscess in rat by using ^99mTc-leukocytes were comparable to those obtained with ¹¹¹In-leukocytes.     

Biological distribution of 99mTc-labeled YIGSR and IKVAV laminin peptides in rodents: 99mTc-IKVAV peptide localizes to the lung

Two laminin-derived peptides containing either YIGSR or IKVAV (single amino acid code) sequences were radiolabeled with ^99mTc and their biological distribution evaluated in rodents. Both ^99mTC-peptides cleared rapidly from the circulation though the kidney, and to a lesser extent, through the liver. ^99mTC-YIGSR peptide did not accumulate in any organ examined in normal, tumored, and emphysemic mice. The ^99mTc-IKVAV peptide localized within 10 min to the lung of normal animals, resulting in lung-to-blood ratios of approximately 23:1. The ^99mTc-IKVAV peptide localized to lung after submicron filtration and after intraperitoneal injection, suggesting that particulates do not play a major role in localization. Pre-incubation of ^99mTc-IKVAV peptide in whole blood decreased lung localization, suggesting that margination of radiolabeled blood cells does not play a major role in the lung localization. When ^99mTc-IKVAV was injected into mice with tumored lungs (melanoma), the lung uptake was markedly increased (up to 20% injected dose higher than control lungs) at all time points examined (10, 30, and 120 min). When ^99mTc-IKVAV was injected into mice with genetic emphysema, the lung uptake was markedly decreased at all time points. The localization of the ^99mTc-IKVAV-containing peptide to the lung is consistent with a receptor-based mechanism.     

Development and Evaluation of a Novel 99mTc-Labeled Annexin A5 for Early Detection of Response to Chemotherapy

^99mTc-HYNIC-annexin A5 can be considered as a benchmark in the field of apoptosis imaging. However, ^99mTc-HYNIC-annexin A5 has characteristics of high uptake and long retention in non-target tissues such as kidney and liver. To minimize this problem, we developed a novel ^99mTc-labeled annexin A5 using a bis(hydroxamamide) derivative [C₃(BHam)₂] as a bifunctional chelating agent, and evaluated its usefulness as an imaging agent for detecting apoptosis. The amino group of C₃(BHam)₂ was converted to a maleimide group, and was coupled to thiol groups of annexin A5 pretreated with 2-iminothiolane. ^99mTc labeling was performed by a ligand exchange reaction with ^99mTc-glucoheptonate. Biodistribution experiments for both ^99mTc-C₃(BHam)₂-annexin A5 and ^99mTc-HYNIC-annexin A5 were performed in normal mice. In addition, in tumor-bearing mice, the relationship between the therapeutic effects of chemotherapy (5-FU) and the tumor accumulation of ^99mTc-C₃(BHam)₂-annexin A5 just after the first treatment of 5-FU was evaluated. ^99mTc-C₃(BHam)₂-annexin A5 was prepared with a radiochemical purity of over 95%. In biodistribution experiments, ^99mTc-C₃(BHam)₂-annexin A5 had a much lower kidney accumulation of radioactivity than ^99mTc-HYNIC-annexin A5. In the organs for metabolism, such as liver and kidney, radioactivity after the injection of ^99mTc-HYNIC-annexin A5 was residual for a long time. On the other hand, radioactivity after the injection of ^99mTc-C₃(BHam)₂-annexin A5 gradually decreased. In therapeutic experiments, tumor growth in the mice treated with 5-FU was significantly inhibited. Accumulation of ^99mTc-C₃(BHam)₂-annexin A5 in tumors significantly increased after 5-FU treatment. The accumulation of radioactivity in tumor correlated positively with the counts of TUNEL-positive cells. These findings suggest that ^99mTc-C₃(BHam)₂-annexin A5 may contribute to the efficient detection of apoptotic tumor response after chemotherapy.     

Preparation and bioevaluation of 99mTc-HYNIC-annexin B1 as a novel radioligand for apoptosis imaging

Annexin B1, a novel Ca²⁺-dependent PS-binding protein, has been shown to have a high affinity for PS exposed on the surface of apoptotic cells. To develop and bioevaluate an annexin B1 based PS-targeting radiotracer, annexin B1 was radiolabeled with ^99mTc using HYNIC as a bifunctional chelator. Binding assays with activated platelets and apoptotic SP2/0 cells were carried out to evaluate the in vitro biological activity of ^99mTc-HYNIC-annexin B1. Biodistribution of this radioligand was studied in normal mice. Dexamethasone-induced murine thymus apoptosis and fas-mediated murine liver apoptosis models were used to investigate the ability of radiolabeled annexin B1 to detect apoptosis in vivo. The labeling procedure yielded a compound with up to 98% radiochemical purity and good in vitro stability. The in vitro binding assays indicated that ^99mTc-HYNIC-annexin B1 retain its PS-binding activity. Biodistribution of the compound in mice showed that ^99mTc-HYNIC-annexin B1 is rapidly cleared from the blood and predominantly accumulates in the kidney. The marked increase in dexamethasone-treated murine thymus uptake and fas-mediated murine liver uptake correlated with histologic evidence of apoptosis. These data suggested that ^99mTc-HYNIC-annexin B1 retain its in vitro and in vivo biological activities. This radiotracer may therefore be useful as a novel radioligand for the noninvasive detecting of PS externalization associated with apoptosis.     

Development of 99mTc-conjugated JS001 antibody for in vivo mapping of PD-1 distribution in murine

Here we reported the development of a novel immuno-SPECT tracer, namely ^99mTc-JS001, to non-invasively image PD-1 expression in mice. The JS001 antibody was directly labeled by the most widely used SPECT radionuclide ^99mTc with a radiochemical yield of 90%, and the specific activity was ≤74 GBq/mmol. After the radiolabeling, ^99mTc-JS001 exhibited a similar immnuoaffinity to PD-1 in vitro. ^99mTc-conjugated JS001 maintained intact in 5% HSA system for 24 h. S180 sarcoma xenograft-bearing Kunming mice and BGC823 gastric cancer orthotopic tumor model were built. Bio-distribution and/or immuno-SPECT studies with ^99mTc-JS001 showed the antibody maintained in the blood, liver, kidneys and tumors at 1.5 ID%/g, 1.4 ID%/g, 2.0 ID%/g and 0.5 ID%/g, respectively. Also, there was a higher uptake in the BGC823 orthotopic tumor than that in the adjunct stomach. These results demonstrated that ^99mTc-JS001 might have capacity to monitor the PD-1 expression in vivo, which might facilitate the anti-PD-1 antibodies treatment in preclinical models.     

99mTc标记BmK CT及其胶质瘤荷瘤裸鼠的SPECT成像研究

目的:通过放射性核素~(99m)Tc标记BmK CT多肽制备靶向胶质瘤的显像剂,探讨~(99m)?Tc-BmK CT用于胶质瘤显像的可行性。方法:采用BmK CT多肽游离的氨基与DTPA酸酐反应得到BmK CT-DTPA,经99m Tc标记后通过柱层析分离纯化制备~(99m)?Tc-BmK CT。测定标记物在PBS溶液和血清中不同时间点放射性化学纯度,评价BmK CT-~(99m)?Tc体外稳定性。新西兰白兔耳缘静脉注射~(99m)Tc-BmK CT进行SPECT显像,观察不同时间点体内的放射性分布。皮下胶质瘤裸鼠经尾静脉注射~(99m)Tc-BmK CT,观察不同时间点肿瘤的摄取情况;注射后4 h处死裸鼠,分离肿瘤和主要器官进行离体SPECT显像,并用勾画感兴趣区法分析相对放射性计数。结果:~(99m)Tc标记BmK CT多肽标记率大于80%,经柱层析分离纯化后放射性化学纯度大于99%。标记物在PBS和血清稳定性良好,6 h内放射性化学纯度均大于95%,12 h内放射性化学纯度大于90%。正常白兔SPECT显像表明~(99m)Tc-BmK CT主要浓聚在肝脏、脾脏和肾脏,软组织持续显影微弱,甲状腺区及胃肠未见核素浓聚;显像剂主要通过泌尿系统排泄,24 h肾脏与肝脏显影接近。胶质瘤裸鼠SPECT显像表明,注射后4 h肿瘤显像清楚,ROI分析结果显示肿瘤/肌肉比4.26±0.25,标记物在肿瘤内代谢缓慢,8 h肿瘤部位仍有较高摄取。结论:本研究成功制备了~(99m)Tc标记BmK CT多肽,标记物主要被肝、脾和肾摄取,经泌尿系统排泄;~(99m)Tc-BmK CT能够在皮下胶质瘤中浓聚,注射后4 h肿瘤显影清晰,瘤内代谢缓慢,有潜力成为一种新型胶质瘤分子探针。     

Measurement of Technetium-99m Sestamibi Signals in Rats Administered a Mitochondrial Uncoupler and in a Rat Model of Heart Failure

Background

Many methods have been used to assess mitochondrial function. Technetium-99m sestamibi (^99mTc-MIBI), a lipophilic cation, is rapidly incorporated into myocardial cells by diffusion and mainly localizes to the mitochondria. The purpose of this study was to investigate whether measurement of ^99mTc-MIBI signals in animal models could be used as a tool to quantify mitochondrial membrane potential at the organ level.

Methods and Results

We analyzed ^99mTc-MIBI signals in Sprague-Dawley (SD) rat hearts perfused with carbonyl cyanide m-chlorophenylhydrazone (CCCP), a mitochondrial uncoupler known to reduce the mitochondrial membrane potential. ^99mTc-MIBI signals could be used to detect changes in the mitochondrial membrane potential with sensitivity comparable to that obtained by two-photon laser microscopy with the cationic probe tetramethylrhodamine ethyl ester (TMRE). We also measured ^99mTc-MIBI signals in the hearts of SD rats administered CCCP (4 mg/kg intraperitoneally) or vehicle. ^99mTc-MIBI signals decreased in rat hearts administered CCCP, and the ATP content, as measured by ³¹P magnetic resonance spectroscopy, decreased simultaneously. Next, we administered ^99mTc-MIBI to Dahl salt-sensitive rats fed a high-salt diet, which leads to hypertension and heart failure. The ^99mTc-MIBI signal per heart tissue weight was inversely correlated with heart weight, cardiac function, and the expression of atrial natriuretic factor, a marker of heart failure, and positively correlated with the accumulation of labeled fatty acid analog. The ^99mTc-MIBI signal per liver tissue weight was lower than that per heart tissue weight.

Conclusion

Measurement of ^99mTc-MIBI signals can be an effective tool for semiquantitative investigation of cardiac mitochondrial membrane potential in the SD rat model by using a chemical to decrease the mitochondrial membrane potential. The ^99mTc-MIBI signal per heart tissue weight was inversely correlated with the severity of heart failure in the Dahl rat model.     

Comparison of 90Y SIRT predicted and delivered absorbed doses using a PSF conversion method

PurposeThe aims of this study were to develop and apply a method to correct for the differences in partial volume effects of pre-therapy Technetium-99 m (^99mTc)-MAA SPECT and post-therapy Yttrium-90 (⁹⁰Y) bremsstrahlung SPECT imaging in selective internal radiation therapy, and to use this method to improve quantitative comparison of predicted and delivered ⁹⁰Y absorbed doses.MethodsThe spatial resolution of ^99mTc SPECT data was converted to that of ⁹⁰Y SPECT data using a function calculated from ^99mTc and ⁹⁰Y point spread functions. This resolution conversion method (RCM) was first applied to ^99mTc and ⁹⁰Y SPECT phantom data to validate the method, and then to clinical data to assess the power of ^99mTc SPECT imaging to predict the therapeutic absorbed dose.ResultsThe maximum difference between absorbed doses to phantom spheres was 178%. This was reduced to 27% after the RCM was applied.The clinical data demonstrated differences within 38% for mean absorbed doses delivered to the normal liver, which were reduced to 20% after application of the RCM. Analysis of clinical data showed that therapeutic absorbed doses delivered to tumours greater than 100 cm³ were predicted to within 52%, although there were differences of up to 210% for smaller tumours, even after the RCM was applied.ConclusionsThe RCM was successfully verified using phantom data. Analysis of the clinical data established that the ^99mTc pre-therapy imaging was predictive of the ⁹⁰Y absorbed dose to the normal liver to within 20%, but had poor predictability for tumours smaller than 100 cm³.     

Technetium-99m-labeled platelets: Comparison of labeling with a new lipid-soluble Sn(II)-mercaptopyridine-N-oxide and 99mTc-HMPAO

Platelets pretinned with a neutral Sn(II)-2-mercaptopyridme-N-oxide (SN-MPO) were labeled with ^99mTc and compared to those labeled with ^99mTc-HMPAO. The conditions of labeling platelets, e.g. concentrations of platelets and Sn(II)-MPO, ^99mTc in ACD-saline or ACD-plasma media, pH and incubation time, were optimized using canine platelets. Moderate labeling efficiency was obtained with 20 μg of tin(II) chloride and 30 min incubation with Sn-MPO and pertechnetate. The viability of labeled platelets was determined by platelet recovery and platelet survival times in Beagle dogs. The labeling efficiency with platelets from 43 mL of blood was 62.8 ± 7.6%. The platelet recovery was 35.7 ± 5.0% and exponential survival time was 34.6 ± 3.1 h compared to 43.3 ± 12.0% and 29.5 ± 3.3 h for ^99mTc-HMPAO-labeled platelets. These values were significantly (P < 0.01) lower than ¹¹¹In-labeled platelets. Biodistribution in dogs indicates lower retention in blood, spleen and liver after some initial ^99mTc excretion in urine. The platelet deposition with ^99mTc platelets (Sn-MPO method) on polyurethane angio-catheters was similar to ^99mTc-HMPAO-labeled platelets. This study indicates that the platelets could be successfully labeled with pertechnetate in a cost-effective manner for the evaluation of thromboembolic complications.     

Preparation,radiolabeling and biodistribution of a new class of bisaminothiol (BAT) ligands as possible imaging agents

In developing new ligands as potential brain and heart perfusion imaging agents two ligands based upon N₂S₂ donor atoms with the biphenyl backbone were synthesized. Biphenyl-2,2′-bis(N-1-amino-2-methyl-propane-2-thiol) (BP-BAT-TM) and biphenyl-2,2′-bis(N-1-amino-2-ethyl-butane-2-thiol) (BP-BAT-TE) form stable, neutral and lipid soluble complexes with [^99mTc]pertechnetate in the presence of tin(II) tartarate as a reducing agent. The [^99mTc]BP-BAT-TM complex penetrates the blood-brain barrier following i.v. injection into rats. Washout from the brain is fast, indicating no retention. The biodistribution of [^99mTc]BP-BAT-TE in rats showed an intitial heart uptake (0.8% /organ, at 2 min) and a slow washout (0.74% at 15 min). No brain uptake was found (0.05%). Significant uptake and retention in liver was observed. An imaging study of [^99mTc]BP-BAT-TE in a monkey showed no brain uptake and a clear indication of liver uptake and gall bladder clearance. These results indicate that this ligand system may be suitable as the basic core structure for the development of new imaging agents. Further studies with structural variations in the biphenyl backbone are warranted to develop new ^99mTc imaging agents for clinical applications.     

99mTc-bioorthogonal click chemistry reagent for in vivo pretargeted imaging

Metal-free click chemistry has become an important tool for pretargeted approaches in the molecular imaging field. The application of bioorthogonal click chemistry between a pretargeted trans-cyclooctene (TCO) derivatized monoclonal antibody (mAb) and a ^99mTc-modified 1,2,4,5-tetrazine for tumor imaging was examined in vitro and in vivo. The HYNIC tetrazine compound was synthesized and structurally characterized, confirming its identity. Radiolabeling studies demonstrated that the HYNIC tetrazine was labeled with ^99mTc at an efficiency of >95% and was radiochemically stable. ^99mTc–HYNIC tetrazine reacted with the TCO–CC49 mAb in vitro demonstrating its selective reactivity. In vivo biodistribution studies revealed non-specific liver and GI uptake due to the hydrophobic property of the compound, however pretargeted SPECT imaging studies demonstrated tumor visualization confirming the success of the cycloaddition reaction in vivo. These results demonstrated the potential of ^99mTc–HYNIC–tetrazine for tumor imaging with pretargeted mAbs.     

Technetium-99m monoclonal antibody fragment (FAb) scintigraphy in the evaluation of small cell lung cancer: a preliminary report

Small cell lung cancer (SCC) has the most rapid growth rate of the four cell types and metastasizes early. Present imaging modalities for staging include chest x-ray, CT, MRI and bone scans. In this preliminary study, we assessed the clinical role of ^99mTc-monoclonal antibody (MOAB) scintigraphy in five patients with histologically proven SCC. Each patient was infused with 20–30 mCi of ^99mTc labeled Fab fragment of MOAB (NR-LU-10, NeoRx, Seattle, Wash.). Total body simultaneous anterior and posterior images were obtained 14–16 h post injection. SPECT images of the chest were obtained through a 360 ° rotation of the gamma camera and recorded on a 62 × 64 × 16 matrix. Images (1.2cm thick) were generated in transaxial, sagittal and coronal views.Fourteen of fifteen chest lesions detected by CT were confirmed by ^99mTc MOAB scintigraphy. Scintigraphy detected one additional chest lesion not seen by CT. Scintigraphy failed to detect a brain lesion (2 cm), a chest lesion, and two adrenal lesions, all of which were seen by CT. In one patient with multiple (more than 10) lesions in the liver, both scintigraphy and CT detected all lesions. Three spine lesions seen on ^99mTc MDP scan and positive for metastasis on MRI concentrated ^99mTc MOAB, but two rib lesions seen on ^99mTc MDP bone scan did not concentrate ^99mTc MOAB. It is concluded from these preliminary results that the potential usefulness of ^99mTc MOAB scintigraphy as a complementary imaging modality in the staging of small cell lung cancer should be investigated further.     

Liposomes for liver scintigraphy

Abstract

Liposomes were prepared by detergent dialysis method and ^99mTc-GHA, a radiopharmaceutical of choice for kidney imaging was entrapped in SUVs of different surface charges. The organ specificity of the agent with and without encapsulation in liposomes was studied in experimental animals. Results obtained show that ^99mTc-GHA entrapped SUV can be targeted to liver. These results were corroborated by scintigraphy studies. The effect of entrapping the agent in SUV on its half-life is discussed.     

A new synthetic method for 99mTc labeled N-Pyridoxyl-5-methyltryptophan as a hepatoma imaging agent

N-Pyridoxyl-5-methyltryptophan (5-PMT) was synthetized by a simplified method using sodium borohydride for the reduction of a Schiff base of pyridoxal and 5-methyltryptophan. Lyophilized kits containing 5-PMT, stannous chloride and l-(+)-ascorbic acid were prepared and labeled to afford ^99mTc-5-PMT with 96% or higher radiochemical purity analysed by two thin-layer chromatographic solvent systems. ^99mTc-5-PMT showed a rapid blood clearance, a faster hepatobiliary transit and a lower renal retention in comparison with ^99mTc-5-EHIDA in rats. Eleven (61%) of 18 patients with histologically confirmed hepatocellular carcinoma showed positive images at 2 to 5 h after i.v. injection. The smallest tumor that could be identified was 2 cm in diameter with the best tumor/liver ratio of 4. In conclusion, ^99mTc-5-PMT synthesized by sodium borohydride reduction shows great promise as a useful hepatoma imaging agent.     

Functionalization of hydroxy compounds with nitrilotriacetic acid for technetium-99m chelation: Excretory properties of the radiolabelled chelates

Substituted monoanilides of nitrilotriacetic acid (NTA) have gained much popularity in recent years as an important class of ligands for technetium-99m (^99mTc) radiopharmaceutical preparations used in liver imaging and function studies. We were interested in investigating the properties of the corresponding ester analogues of this important class of ligands and for this study cyclohexanol was selected as a hydroxy component, which on condensation with nitrilotriacetic acid in the presence of acetic anhydride, furnished the monoester, N-cyclohexyloxycarbonylmethyl iminodiacetic acid 4 and the corresponding diester 5. Phenol on similar condensation produced mainly the diester, N, N-di(phenyloxycarbonylmethyl) aminoacetic acid 2, with traces of the corresponding monoester 7. A reinvestigation of the well known condensation reaction of aniline with nitrilotriacetic acid revealed that in addition to the reported monoanilide, N-phenylcarbamoylmethyl imino diacetic acid 3, the corresponding dianilide 6 was also produced in appreciable amount. The ester ligands 2, 4, 5 after ^99mTc chelation exhibited good in vitro and in vivo stabilities. The biodistribution characteristics of these radiolabelled esters and amides were very similar showing thereby that esterification with NTA could be an effective method for converting alcohols to ^99mTc-radiopharmaceuticals without generating any unusual properties because of the ester linkage. Residual radiopharmaceutical concentration after i.v. administration of these amide and ester ^99mTc chelates at 30 min in blood, urine, liver, kidney and intestine were correlated with their lipophilicities and during this correlation it was observed that in addition to lipophilicity the anionic strength of these chelates is also an important determinant in governing their biodistribution. The ester ligand 4 after ^99mTc chelation showed ultrafast hepatobiliary kinetics and was therefore compared in a rabbit model with a standard hepatobiliary radiopharmaceutical ^99mTc-N-(p-butylphenylcarbamoyl methyl) iminodiacetic acid (^99mTc-BIDA) by γ-camera scintigraphy to investigate the potential of the former for clinical studies.     

Evaluation of a DMSA kit for instant preparation of 99mTc(V)—DMSA for tumour and metastasis scintigraphy

A kit has been developed to instantly prepare ^99mTc(V)—DMSA. The freeze-dried kit consisting of DMSA, stannous chloride and ascorbic acid in appropriate proportions, produces quality ^99mTc(V)—DMSA when mixed with 0.2 mL of 3.5% NaHCO₃ solution and 2–4 mL of [^99mTc] pertechnetate. The radiopharmaceutical characterized by chromatography with ITLC-SG in 0.9% saline and horizontal paper electrophoresis in 50 mM vernol buffer, pH 8.6, at a potential gradient of 15 V/cm showed a different mobility with respect to ^99mTc(III)-DMSA, a known agent for kidney imaging. The new agent exhibited less plasma protein binding as compared to that of ^99mTc(III)-DMSA. Biodistribution of the pentavalent DMSA in mouse demonstrated greater uptake in bone and muscle and lower uptake in liver and kidney with respect to trivalent DMSA. The soft tissue tumour specificity and its suitability for tumour scintigraphy was apparent from the scintigrams of mammary carcinoma in a C₃H Jax mouse and medullary carcinoma in a patient. Brain metastatic lesions were also visible in a breast carcinoma patient after administering him with the agent.     

Capillary electrophoresis of technetium radiopharmaceuticals

Diagnostically used ^99mTc kit radiopharmaceuticals were analyzed using capillary zone electrophoresis with radioactivity detection: ^99mTc-bis(bis(2-ethyloxyethyl)phosphino)ethane (^99mTc-Myoview, ^99mTc-Tetrofosmin), ^99mTc-trans(1,2-bis(dehydro-2,2,5,5,-tetramethyl-3-furanone-4-methylene-amino)ethane)-tris(3-methoxy-1-propyl)phosphine) (^99mTc-Technescan Q12, ^99mTc-Furifosmin), ^99mTc-methoxyisobutylisonitrile (^99mTc-MIBI), ^99mTc- , -ethylenecysteine diethylester dimer (^99mTc-ECD), ^99mTc-d,l-hexamethylene propyleneamine oxime (^99mTc-HMPAO), ^99mTc-diethylenetriaminepentaacetic acid (^99mTc-DTPA), ^99mTc-ethylene hepatobiliary iminodiacetic acid (^99mTc-EHIDA), ^99mTc- , -ethylenecysteine dimer (^99mTc-EC), ^99mTc-mercaptoacetylglycylglycylglycine (^99mTc-MAG₃), ^99mTc-dimercaptosuccinic acid (^99mTc-DMSA), ^99mTc-methylene diphosphonate (^99mTc-MDP) and ^99mNaTcO₄. A pressure-driven capillary zone electrophoresis was employed to detect small anions of high electrophoretic mobility and cations within one run. Effective ^99mTc complex charges could be determined by a neutral internal standard. All complexes showed the expected electrophoretic behaviours in view of their charges. Pure products were obtained for the majority of the studied complexes. In the case of ^99mTc-Q12, ^99mTc-EHIDA and ^99mTc-MDP, complex mixtures were detected. The high potential of CE for the analysis of ^99mTc radiopharmaceuticals could be shown.     

A simple method for producing a technetium-99m-labeled liposome which is stable In Vivo

A new method for labeling preformed liposomes with technetium-99m (^99mTc) has been developed which is simple to perform and stable in vivo. Previous ^99mTc-liposome labels have had variable labeling efficiencies and stability. This method consistently achieves high labeling efficiencies (> 90%) with excellent stability. A commercially available radiopharmaceutical kit—hexamethylpropyleneamine oxime (HM-PAO)—is reconstituted with ^99mTcO^−₄ and then incubated with preformed liposomes that encapsulate glutathione. The incubation takes only 30 min at room temperature. Liposomes that co-encapsulate other proteins such as hemoglobin or albumin, in addition to glutathione, also label with high efficiency. Both in vitro and in vivo studies indicate good stability of this label. Rabbit images show significant spleen and liver uptake at 2 and 20 h after liposome infusion without visualization of thyroid, stomach or bladder activity.This labeling method can be used to study the biodistribution of a wide variety of liposome preparations that are being tested as novel drug delivery systems. This method of labeling liposomes with ^99mTc may also have applications in diagnostic imaging.     

Capillary electrophoresis of 99mtechnetium radiopharmaceuticals

Diagnostically used ^99mTc kit radiopharmaceuticals were analyzed using capillary zone electrophoresis with radioactivity detection: ^99mTc-bis(bis(2-ethyloxyethyl)phosphino)ethane (^99mTc-Myoview, ^99mTc-Tetrofosmin), ^99mTc-trans(1,2-bis(dehydro-2,2,5,5,-tetramethyl-3-furanone-4-methylene-amino)ethane)-tris(3-methoxy-1-propyl)phosphine) (^99mTc-Technescan Q12, ^99mTc-Furifosmin), ^99mTc-methoxyisobutylisonitrile (^99mTc-MIBI), ^99mTc-l,l-ethylenecysteine diethylester dimer (^99mTc-ECD), ^99mTc-d,l-hexamethylene propyleneamine oxime (^99mTc-HMPAO), ^99mTc-diethylenetriaminepentaacetic acid (^99mTc-DTPA), ^99mTc-ethylene hepatobiliary iminodiacetic acid (^99mTc-EHIDA), ^99mTc-l,l-ethylenecysteine dimer (^99mTc-EC), ^99mTc-mercaptoacetylglycylglycylglycine (^99mTc-MAG₃), ^99mTc-dimercaptosuccinic acid (^99mTc-DMSA), ^99mTc-methylene diphosphonate (^99mTc-MDP) and ^99mNaTcO₄. A pressure-driven capillary zone electrophoresis was employed to detect small anions of high electrophoretic mobility and cations within one run. Effective ^99mTc complex charges could be determined by a neutral internal standard. All complexes showed the expected electrophoretic behaviours in view of their charges. Pure products were obtained for the majority of the studied complexes. In the case of ^99mTc-Q12, ^99mTc-EHIDA and ^99mTc-MDP, complex mixtures were detected. The high potential of CE for the analysis of ^99mTc radiopharmaceuticals could be shown.     

Rat hepatocytes bind to synthetic galactoside surfaces via a patch of asialoglycoprotein receptors

The binding of rat hepatocytes to flat polyacrylamide surfaces containing galactose is sugar-specific, requires Ca+2, and occurs only above a critical concentration of sugar in the substratum [Weigel et al., 1979, J. Biol. Chem., 254, 10,830). Binding is completely inhibited by asialo-orosomucoid but not by orosomucoid or asialo- agalacto-orosomucoid, suggesting that cell binding is mediated by asialoglycoprotein receptors. Asialo-orosomucoid was labeled with fluorescein isothiocyanate and used as a direct fluorescent probe to monitor the distribution of cell surface asialoglycoprotein receptors before and after hepatocyte binding to galactoside or control substrata. Cells bound at 37 degrees C were de-adhered at 4 degrees C using the Ca+2 chelator EGTA. The released cells were then stained with fluorescein-asialo-orosomucoid, fixed, washed, and examined by fluorescence microscopy. On freshly isolated cells before binding, the distribution of asialoglycoprotein receptors appears diffuse and nonclustered. However, more than half of the cells released intact from a galactoside surface had a single large (4 micrometer2) fluorescent patch. The receptor patch cannot be detected on cells while they are bound to a galactoside surface but rather only on released cells, indicating that the cell-substratum junction is the site of the receptor patch. No asialoglycoprotein receptor patches (less than or equal to 1%) were observed on cells that were incubated on, but did not bind to, an underivatized polyacrylamide surface or to a surface with a galactose concentration below the critical concentration for binding. Furthermore, no receptor patches were present on cells that had bound to and were subsequently released from substrata that did not contain galactose, including glass, tissue culture plastic, nontissue culture plastic, and collagen. The distribution of asialoglycoprotein receptors is preserved at 4 degrees C because at 37 degrees C the patches disappear with a half-life of approximately 2.6 min. The results directly demonstrate that a large cluster of asialoglycoprotein receptors mediates the binding of rat hepatocytes to a galactoside surface.