Efficient one-step direct labelling of recombinant antibodies with technetium-99m

High-affinity bacterially expressed antibody fragments can nowadays be cloned from established hybridomas or, more conveniently, isolated directly from antibody libraries displayed on filamentous phage. Such antibodies can be tagged with C-terminal peptide tags containing one cysteine residue, which represents a convenient functionalisation site for a number of applications, including technetium-99m labelling. Here we describe a simple one-step method for^99m-Tc labelling of cysteine-tagged recombinant antibodies with more than 50% radionuclide incorporation. The labelled antibodies displayed full retention of immuoreactivity and good stability.     

Technetium-99m and indium-111 double labelling of granulocytes for kinetic and clinical studies

A new technique of labelling granulocytes with both technetium-99m hexamethylpropylene amine oxime (HMPAO) and indium-111 in a single protocol was developed in order to exploit the advantages of each radiolabel in clinical and investigative studies. Fourteen patients were included in this prospective study. Granulocytes were labelled with both¹¹¹In-tropolonate and^99mTc-HMPAO. In vitro shape change assay and in vivo distribution and recovery studies were performed to assess the activation of and damage to these cells due to the labelling procedure. The comparative kinetics of¹¹¹In and^99mTc in the blood, liver, spleen, and bone marrow were studied by blood sampling and dual radionuclide imaging early (1 h) and late (24 h) after injection. The functional integrity of the double-labelled granulocytes and the feasibility of the technique were investigated in 14 patients with a painful prosthetic hip due to causes other than infection. The efficiency of double labelling was 63% (SD 14%) for¹¹¹In and 39% (SD 12%) for^99mTc-HMPAO. In vitro granulocyte activation and ex vivo recovery values were comparable to those from single radionuclide labelling. No artefactual granulocyte sequestration was seen in the lungs or liver. The radioactivity was distributed between the liver, spleen and bone marrow and, to a lesser extent, the lung. Early^99mTc counts in the liver, spleen and bone marrow, in relation to background, were significantly higher than¹¹¹In counts while the reverse was seen in late images. Furthermore, circulating free^99mTc was significantly higher than free¹¹¹In at 24 h. Organ^99mTc counts, expressed in relation to the activity in early images, decreased in the spleen, increased in the liver and remained unchanged in bone marrow, whereas¹¹¹In counts increased in the bone marrow and liver, and decreased in the spleen. Granulocytes can be labelled with both¹¹¹In and^99mTc-HMPAO in a single protocol without crosschelation, cellular activation or damage. By favourably exploiting their kinetics for early and late imaging, double-labelled granulocytes may be useful in several clinical and investigative situations.     

High-quality <Superscript>124</Superscript>I-labelled monoclonal antibodies for use as PET scouting agents prior to <Superscript>131</Superscript>I-radioimmunotherapy

Purpose Monoclonal antibodies (MAbs) labelled with ¹²⁴I are an attractive option for quantitative imaging with positron emission tomography (PET) in a scouting procedure prior to ¹³¹I-radioimmunotherapy (¹³¹I-RIT). In this study, three important items in the labelling of MAbs with ¹²⁴I were introduced to obtain optimal and reproducible product quality: restoration of radiation-induced inorganic deterioration of the starting ¹²⁴I solution, radiation protection during and after ¹²⁴I labelling, and synchronisation of the I/MAb molar ratio.Methods A new method was applied, using an NaIO₃/NaI carrier mix, realising in one step >90% restoration of deteriorated ¹²⁴I into the iodide form and chemical control over the I/MAb molar ratio. Chimeric MAb (cMAb) U36 and the murine MAbs 425 and E48 were labelled with ¹²⁴I using the so-called Iodogen-coated MAb method, as this method provides optimal quality conjugates under challenging radiation conditions. As a standardising condition, NaIO₃/NaI carrier mix was added at a stoichiometric I/MAb molar ratio of 0.9. For comparison, MAbs were labelled with ¹³¹I and with a mixture of ¹²⁴I, ¹²³I, ¹²⁶I and ¹³⁰I.Results Labelling with ¹²⁴I in this setting resulted in overall yields of >70%, a radiochemical purity of >95%, and preservation of MAb integrity and immunoreactivity, including at the patient dose level (85 MBq). No significant quality differences were observed when compared with ¹³¹I products, while the iodine isotope mixture gave exactly the same labelling efficiency for each of the isotopes, excluding a different chemical reactivity of ¹²⁴I-iodide. The scouting performance of ¹²⁴I-cMAb U36 labelled at the patient dose level was evaluated in biodistribution studies upon co-injection with ¹³¹I-labelled cMAb U36, and by PET imaging in nude mice bearing the head and neck cancer xenograft line HNX-OE. ¹²⁴I-cMAb and ¹³¹I-cMAb U36 labelled with a synchronised I/MAb molar ratio gave fully concordant tissue uptake values. Selective tumour uptake was confirmed with immuno-PET, revealing visualisation of 15 out of 15 tumours.Conclusion These results pave the way for renewed evaluation of the potential of ¹²⁴I-immuno-PET for clinical applications.     

Tc-99m-HMPAO labelled human platelets: in vitro and in vivo results

The lipophilic ^99mTc-HMPAO complex can be used for labelling platelets as well as granulocytes. Platelets were isolated according to standard isolation procedures for the evaluation of the optimal labelling parameters. The labelling efficiency (%) depends on incubation temperature (22° C: 40%; 37° C: 50%), incubation time (3 min: 20%, 25 min: 55%) and the incubation medium (plasma: 40%; saline 50%). The 60 min ^99mTc elution, out of the platelets ranged around 8%. The platelet recovery used as a quality control parameter is around 25%±4% and is stable for at least 240 min. The high elution rate out of the platelets leads to renal excretion of the label and hence to significant kidney and bladder activity. Intestinal excretion of the label can also be frequently demonstrated. Fresh thrombotic lesions can normally be detected 4 h after reinjection of the labelled platelets, and in some patients as early as 1 h after reinjection. In conclusion, ^99mTc-HMPAO seems to be a promising platelet label for imaging thrombotic lesions but not for platelet survival studies, because of the short physical half life of ^99mTc.Supported by the Deutsche Forschungsgemeinschaft (BE 1054/1–2)     

In vitro comparison of HMPAO and gentisic acid for labelling leukocytes with99mTc

Leukocytes can be labelled with^99mTc using HMPAO and gentisic acid methods. We compared the two methods with respect to labelling efficiency on mixed leukocytes and isolated polymorphonuclear (PMN) and mononuclear (MN) cells, and the in vitro stability of the label. HMPAO produced approximately 70% labelling efficiency on mixed or PMN cells and the label was stable in saline or plasma. Labelling efficiency on MN was only 14% and was less stable. Gentisic acid produced a labelling efficiency of 52% on PMN and 35% on MN, both were stable in saline but less stable in plasma. In conclusion, HMPAO produces higher labelling efficiency and the label shows greater in vitro stability in plasma. However, gentisic acid is much less expensive to use, allows labelling of MN cells, and should result in more favourable microdosimetry. Preliminary clinical results suggest that gentisic acid is equivalent to HMPAO but has the advantage of being much cheaper.     

New fast preparation of 123I labelled radiopharmaceuticals

A new fast kit preparation of ¹²³I labelled radio-pharmaceuticals such as IMP, HIPDM, MIBG and Hippuran is proposed. A radiochemical yield >99% is obtained at 100°C within 10–30 min. The new labelling procedure is based on the nucleophilic exchange in presence of Cu(I) and an excess of reducing agents. The four kit prepared ¹²³I-radiopharmaceuticals have been used with success in clinical studies involving 400 patients. The proposed method is also compared with earlier described methods which, yielding labelled side products and ¹²³I₂, do not fulfill the requirements for kit labelling.     

Preparation and evaluation of the rhenium-188-labelled anti-NCA antigen monoclonal antibody BW 250/183 for radioimmunotherapy of leukaemia

Anti-NCA antigen antibody BW 250/183 (Anti-Granulocyte) localizes more than 50% of injected antibody dose to the bone marrow. Therefore, this antibody is promising for adjuvant conditioning radioimmunotherapy of bone marrow before bone marrow transplantation. To examine its potential use for radioimmunotherapy, we developed an efficient and reproducible technical protocol for labelling anti-NCA antigen antibody BW 250/183 with generator-produced rhenium-188, aiming at both high radiochemical yield and high specific activity. ¹⁸⁸Re-labelled BW 250/183 antibody was used in 12 patients with advanced leukaemia. Labelling of BW 250/183 with ¹⁸⁸Re was accomplished by the direct radiolabelling method using tris-(2-carboxyethyl) phosphine (TCEP) as the reducing agent. Twelve patients with recurrent acute or chronic leukaemia were treated with activities of 6.5–12.4 GBq of ¹⁸⁸Re-labelled BW 250/183. Standard gamma camera scintigraphy was used to evaluate the biodistribution, and a region of interest analysis together with the MIRDOSE 3.1 software was applied to determine the radiation doses to relevant tissues. The ¹⁸⁸Re-BW 250/183 antibody was labelled in high radiochemical yield, with high radiochemical purity (94%±3%) and specific activity (5.55–7.4 GBq/mg) within 1 h. The preliminary biodistribution studies showed persistent uptake of ¹⁸⁸Re-BW 250/183 in bone marrow. The radiation absorbed doses (mGy/MBq) delivered to the total body, red marrow, liver, spleen and kidneys were 0.13±0.02, 1.45±0.71, 0.43±0.21, 1.32±0.99 and 0.71±0.17, respectively. TCEP reduction enabled the direct, fast and effective labelling of the monoclonal antibody BW 250/183 with ¹⁸⁸Re. Preliminary clinical results suggest delivery of a significant radiation dose to bone marrow and thus the potential for adjuvant conditioning therapy before BMT. Received 6 January and in revised form 10 May 1999     

Technetium-99m labelled LDL as a tracer for quantitative LDL scintigraphy

The goal of the present study was to optimize technetium-99m labelling of low-density lipoprotein (LDL) and to investigate the in vitro and in vivo properties of the tracer to determine whether its application for quantitative scintigraphy of hepatic LDL receptor activity is feasible. LDL labelled with iodine-125 by the iodine monochloride method was used as a reference tracer. Comparison of different assessments of radiochemical purity [trichloro-acetic acid precipitation (%ppTCA), paper chromatography, size-exclusion chromatography and chloroform-methanol extraction] exhibited %ppTCA to be superior as a parameter of tracer quality. In spite of a high radiochemical purity immediately after labelling, modifications of ^99mTc labelling of LDL did not overcome the poor long-term stability of the tracer. Subsequent dialysis in phosphate buffer over about 3 h sufficiently increased the long-term stability in vitro and in vivo. The competitive recognition of dialysed ^99mTc-LDL and ¹²⁵I-LDL with native LDL by high-affinity binding sites was demonstrated in human hepatoma cells (HepG2) and human fibroblasts. Biodistribution data of simultaneously injected ^99mTc-LDL and ¹²⁵I-LDL in New Zealand White rabbits showed a high uptake of both tracers in tissues with high LDL receptor activity, yet ^99mTc-LDL uptake exceeded ¹²⁵I-LDL uptake by two- to sevenfold. In contrast to ¹²⁵I-LDL, ^99mTc-LDL showed a higher unspecific uptake into the bone marrow and the spleen, suggesting an additional uptake mechanism probably via the scavenger pathway. Curve deconvolution of plasma clearance in five female New Zealand White rabbits and five male hyperlipidaemic patients again showed a marginally different biokinetic behaviour of ^99mTc-LDL and ¹²⁵I-LDL. It is concluded that dialysis of ^99mTc-LDL substantially increases long-term stability, which is essential for quantification purposes, and that the dialysed tracer has retained its biological integrity as it is recognized by the LDL receptor in vitro. It remains to be determined to what extent the estimation of hepatic LDL receptor activity by quantitative ^99mTc-LDL scintigraphy is influenced by the different biokinetic behaviour of ^99mTc-LDL and ¹²⁵I-LDL.     

188Re直接标记octreotide的方法学

188Re标记octreotide可用直接标记法或间接标记法。直接标记法包括预锡化法和分步还原法，分步还原法需先用还原剂还原octreotide，然后用SnCl2还原188ReO4-进行直接标记；预锡化法则直接以SnCl2还原octreotide和188ReO4-，打开octreotide分子内双硫键，使188Re直接标记octreotide。预锡化直接标记法简便快速，能够得到较高的放化纯度，无须进一步纯化，适合用于制备药盒。     

Biological consequences of the heterogeneous irradiation of lymphocytes during technetium-99m hexamethylpropylene amine oxime white blood cell labelling

Technetium-99m hexamethylpropylene amine oxime (^99mTc-HMPAO) labelling of white blood cells, routinely used for the detection of infection, results in the incorporation of radioactivity by polymorphonuclear leucocytes and also lymphocytes and can induce cell lesions in the latter case. The aim of this study was therefore to acquire data on the morphological and functional status of labelled lymphocytes present in the ^99mTc-HMPAO leucocyte mixture and to determine the cellular consequences of labelling. The mean radioactivity associated with lymphocytes was 325±10.8 kBq/10⁶ lymphocytes under standard labelling conditions. Microautoradiographic studies showed that labelling was heterogeneous (4% intensely labelled cells), which prevented calculation of the mean absorbed dose. The frequency of chromosomal aberrations (dicentrics and rings) in the labelled lymphocytes for 380 kBq/10⁶ cells was 1.08±0.09 but no abnormality was observed in the unlabelled control lymphocytes. The plating efficiency of labelled lymphocytes was reduced, as compared with that for control cells, but some lymphocytes were still able to form clones and were still ”alive” by radiobiological definition. It is therefore suggested that lymphocytes should be removed from ^99mTc-HMPAO cell preparations before administration to patients. Received 9 March and in revised form 1 June 1998     

Anomalies in reduction-mediated technetium-99m labelling of monoclonal antibodies

A reduction-mediated technetium-99m labelling method has been evaluated with a range of tumour-specific monoclonal antibodies. Antibodies reduced with 2-mercaptoethanol (2-ME) had free sulphydryl groups, but their number was much higher than could be accounted for by only limited intra-chain reduction of disulphide bonds. Reduced antibody could be labelled efficiently with ^99mTc using an methylene diphosphonate (MDP) bone-scanning kit, although this seemed to depend on the presence of residual 2-ME in the preparations. With a carcinoembryonic antigen (CEA)-specific antibody, immunoreactivity of labelled antibody was confirmed, and after injection into nude with CEA-producing xenografts there was localisation into the tumours. Sephacryl S300 gel filtration showed the radiolabel eluting at a single discrete peak at the expected 150 kDa. However, examination of all of the labelled antibodies by polyacrylamide gel electrophoresis (PAGE) and autoradiography showed the presence of a large number of radiolabelled low molecular weight degradation products of the antibodies. These degradation products seemed to be formed in previously reduced antibodies during processing for PAGE, indicating some fragility of the reduced antibody. Offprint requests to: M.V. Pimm     

99mTc-labelling of leucocytes with 99mTc-DPO: a complex developed for myocardial imaging

The lipophilic ^99mTc-DPO complex, developed as a myocardial imaging radiopharmaceutical, was used to label leucocytes. After an incubation of 0.1 ml ^99mTc-DPO (8 g DMPE*2HCl) with mixed leucocytes in plasma, the labelling efficiency was over 70%. During incubation in 5 ml plasma, a loss of activity was found between 20% (1 h) and 35% (3 h) caused by elution. Disturbances of cell viability could not be found with the help of the chemiluminescence test. The in vivo recovery was determined in three dogs and was 45%–50% (0.5 h), 30%–36% (1 h), and 18%–24% (3 h). Autologous ^99mTc-DPO-leucocytes were used on seven patients with suspected osteomyelitis, there were four true negative and three true positive results. The target/nontarget ratio determined by ROI in the positive cases was 1.8 to 2.5 at 3 h after injection.     

Uptake of technetium-99m hexamethylpropylene amine oxime labelled white cells in lymph nodes involved in non-Hodgkin's lymphoma

Radiolabelled white cell scanning is widely used to detect the presence of infection. We present a case of non-Hodgkin's lymphoma manifesting with signs and symptoms suggestive of infection, in which a technetium-99m hexamethylpropylene amine oxime (^99mTc-HMPAO) white cell scan demonstrated high uptake in lymph nodes involved by lymphoma. Differential cell analysis showed preferential lymphocyte labelling. The classification and management of the disease were changed accordingly. Our findings suggest that a future role for^99mTc-HMPAO labelled white cells in the assessment of disease activity of lymphoma should be investigated.     

Technetium-99m labelled hydrazinonicotinamido human non-specific polyclonal immunoglobulin G for detection of infectious foci: a comparison with two other technetium-labelled immunoglobulin preparations

Recently a new linker — hydrazinonicotinate (HYNIC) — was introduced for labelling of proteins and peptides with technetium-99m. HYNIC and other linkers have been used for labelling of human non-specific polyclonal immunoglobulin G (hIgG) with^99mTc for the detection of infections. In this study we compared the tissue distribution of three different^99mTc-hIgG preparations in groups of five Wistar rats with a focal intramuscular infection withStaphylococcus aureus. We compared^99mTc-HYNIC-hIgG with^99mTc-hIgG labelled via the so-called Schwarz method (reduction of disulphide bonds) and with the^99mTc-labelled commercially available Technescan-HIG. Unlike the HYNIC linker, in the two other labelling methods free sulph-hydryl groups are involved in the binding of^99mTc. High-performance liquid chromatography analysis of the labelled preparations and of plasma samples revealed aggregate or polymer formation in all three agents; this was least pronounced in the product labelled by means of the Schwarz method. The tested preparations did not show signs of degradation in vitro. The difference in linker chemistry was reflected in the tissue distribution. Thus the biodistribution of^99mTc-HYNIC-hIgG was significantly different from the distribution of the two other preparations: abscess (1.4%±0.2%ID/g), muscle, liver, spleen, plasma, lung, bone marrow, and small intestine concentrations were higher at 24 h p.i.; kidney uptake (1.19%±0.08%ID/g) was significantly lower. The abscess-to-plasma and the abscess-to-muscle ratios (0.5 and 11, respectively), however, were in the same range for the three preparations tested. Quantitative analysis of the scintigraphs revealed that the total body clearance of^99mTc-HYNIC-hIgG was significantly slower than for the other agents. The abscess uptake of^99mTc-HYNIC-hIgG as a percentage of the remaining body activity was significantly higher. Based on its high abscess uptake, its low uptake in the kidneys and the high percentage of its abscess uptake in relation to the remaining body activity, we conclude that^99mTc-HYNIC-hIgG seems superior to the two other preparations tested for the detection of infections.     

Radioimmuno targetting (99m)technetium labeled anti-epidermal growth factor receptor monoclonal antibodies in experimental tumor models.

AIM: Monoclonal antibodies (MAb) directed at the extra cellular domain (ECD) of epidermal growth factor receptor (EGFR) offer a promising strategy for diagnosis and therapy of cancers that over-express EGFR. Radiolabelled MAbs against cell surface antigens have improved in vivo tumor diagnosis and treatment. EGFR over-expression has been reported in a wide range of carcinomas especially of the head and neck, breast, etc., and is associated with poor prognosis and resistance to therapy. CIBCNSH3 is a murine MAb generated to the ECD of EGFR in our laboratory and has been extensively characterized and has proven antitumor activity. The tumor targeting potential of (99m)Tc labelled CIBCNSH3 in an experimental tumor model is discussed in this paper. METHODS: A431, an epidermoid carcinoma cell line with overexpression of EGFR, SUDHLH, a lymphoma cell line was used to induce xenografts in inbred adult female BALB/C mice and used for the study. A reduction mediated method of (99m)Tc labelling was adopted to label the MAb. Scintiscan pictures were taken at different time intervals after i.v. administration of the (99m)Tc labelled MAb using a gamma camera and results were correlated with those of biodistribution studies. RESULTS: Immunoscan pictures taken at different time periods showed high uptake of the radioimmunoconjugate by the tumor providing clear tumor images and no uptake in control animals with lymphoma xenografts. Results of scan pictures correlated well with the biodistribution studies. CONCLUSION: The radioimmunoconjugate (99m)Tc-CIBCNSH3 appears to be a promising tool in identifying any early recurrence and micro-metastasis of lesions that overexpress EGFR.     

Technetium-99m labelling of the anti-tumour antibody PR1A3 by photoactivation

Irradiation of antibody with ultraviolet light leads to reduction of disulphide bonds. Thus irradiation can be used to generate free thiols prior to direct labelling of antibody with technetium-99m, and has a potential advantage over methods using chemical reducing agents such as mercaptoethanol or tin, in that no purification step is needed to remove excess reducing agent. We have used the photoactivation method developed by Sykes et al. to label the anti-tumour antibody PR1A3 with^99mTc. The antibody was irradiated at 300 nm using a Rayonet photochemical reactor with eight RMR3000 lamps. In a typical experiment, the antibody solution was injected into a nitrogen-filled borosilicate glass vial and purged with nitrogen. A degassed solution containing stannous fluoride and methylene diphosphonate was then added to the antibody and the vial was irradiated. Following the irradiation, [^99mTc]pertechnetate was injected into the vial and the reaction mixture was incubated for 30 min at room temperature before being analysed by size-exclusion high-pressure liquid chromatography and instant thin-layer chromatography. Labelling yields greater than 95% were obtained using antibody concentrations ranging from 0.5 mg/ml to 5 mg/ml. Irradiation times as short as 5 min and tin to antibody ratios in the range between 11 and 32 µg tin per mg antibody gave high labelling yields. Labelling yields greater than 95% were obtained after storage of the photoactivated antibody at –70° C for several weeks. The stability of the^99mTc-labelled photoactivated PR1A3 was similar to that of^99mTc-labelled mercaptoethanol-reduced PR1A3. The mean immunoreactive fraction was 77% for the photoactivation-labelled PR1A3, compared to 93% for PR1A3 labelled by mercaptoethanol reduction. Biodistribution studies were carried out using^99mTc-photoactivation-labelled PR1A3 or PR1A3 labelled by mercaptoethanol reduction in Balb/c mice and in nude mice with MKN-45 human tumour xenografts. There was no significant difference in tumour uptake between the mice that received photoactivated PR1A3 and those that received mercaptoethanol-reduced PR1A3. There was also no significant difference in uptake in most organs in Balb/c mice; however, the photoactivated antibody cleared more rapidly from the blood, and whole-body clearance was also faster for the photoactivated PR1A3. In conclusion, the photoactivation technique provides a very convenient one-pot method for labelling antibodies with^99mTc.     

Labelling of leucocytes with colloidal tech netium-99m-SnF2: an investigation of the labelling process by autoradiography

Autoradiography of smears and frozen sections of labelled cell suspensions was used to study the distribution of radioactivity in and among blood cells labelled in either whole blood or leucocyte-rich plasma (LRP) with technetium-99m-SnF₂ colloid. The tracer proved selective for neutrophils: the labelling probability (relative to that for erythrocytes) for each cell type in LRP (mean of five samples) was: neutrophils, 9.4; lymphocytes, 3.7; monocytes, 3.0; eosinophils 1.4; erythrocytes, 1.0. When labelling was carried out in whole blood (five samples), 74.5%±8.3% of the cell-bound radioactivity was bound to erythrocytes, 13.6%±6.5% to neutrophils, and 11.9%±2.1% to lymphocytes, whereas in LRP (in which the leucocytes were only slightly out-numbered by erythrocytes), 76.5%±14.9% of radioactivity was neutrophil bound. Labelled cells in smear autoradiographs exhibited two distinct silver grain patterns, diffuse, consistent with an intracellular radioactive particle (in neutrophils), and focal, consistent with a cell surface-adhering particle in direct contact with the emulsion (in other leucocyte types and erythrocytes). The phagocytic inhibitor cytochalasin B neither reduced the proportion of labelled neutrophils nor altered the labelling pattern. Neutrophils were able to scavenge radioactivity from the surface of erythrocytes. It is concluded that neutrophils bind^99mTc-SnF₂ intracellularly by phagocytosis, with high affinity; other cells become labelled at the cell surface reversibly and with lower affinity. This selectivity is high enough to permit predominantly leucocyte labelling in LRP but not in whole blood.     

Intraindividual comparison of 99mTc-labelled anti-SSEA-1 antigranulocyte antibody and 99mTc-HMPAO labelled white blood cells for the imaging of infection

Technetium-99m labelled antigranulocyte antibodies are ready to use and are sensitive and specific in the diagnosis of infectious diseases. ^99mTc-SSEA antigranulocyte antibodies have a very high affinity constant (K _d 10^–12 M) for human neutrophils (PMNs), and excellent imaging qualities with high target/background ratios. The aim of this study was to compare the diagnostic accuracy of the ^99mTc-anti-SSEA-1 monoclonal antibody (Mab) with that of ^99mTc-hexamethylpropylene amine oxime (HMPAO)-labelled white blood cells (WBCs). To this end, 17 patients with 23 proven infectious foci were examined with 555 MBq ^99mTc-anti-SSEA-1 MAb and with 370 MBq ^99mTc-HMPAO labelled autologous leucocytes within a period of 7 days. All the infections were confirmed by culture, biopsy, surgery and follow-up. Whole-body images and planar spot views with the antibody were performed at 1-h, 4-h and 24-h post injection; the biodistribution of the antibody was quantified, absorbed radiation doses were calculated and the diagnostic results were compared with the ^99mTc-HMPAO WBC images. Human anti-mouse antibody (HAMA) evaluation was performed in all patients before and 3 months after antibody injection. Blood was drawn at different times after ^99mTc-anti-SSEA-1 MAb injection to determine the amount of granulocyte-associated radioactivity and to calculate recovery. ^99mTc-anti-SSEA-1 MAb scintigraphy detected all 23 lesions, while 21 were detected with ^99mTc-HMPAO WBC scan. In this small group of patients, the sensitivity and specificity of ^99mTc-anti-SSEA-1 MAb scintigraphy were 95% and 96% respectively, as compared with 91% and 82% respectively for ^99mTc-HMPAO WBC scan. An increasing uptake of the injected activity in the lesion at different time points was indicative of high affinity and of specific PMN binding.There was no HAMA formation. In four of five patients investigated, a transient mild leukopenia was found at 15 min p.i.. There was increased uptake of the antibody in liver and spleen and normal uptake in kidneys and bone marrow.The estimated radiation doses for the whole body and the red bone marrow were 1.1×10^–2 cGy/37 MBq and 5.3×10^–2 cGy/37 MBq, respectively. The activity associated to the PMNs in vivo was 33.5%, 30.6%, 21.3% and 9% at 5, 15, 30 and 45 min. post-injection, respectively. It is councluded that use of ^99mTc-anti-SSEA-1 antigranulocyte antibodies demonstrates promising results comparable to those obtained with ^99mTc-labelled autologous WBCs. The ^99mTc-labelled MAb is ready to use, has excellent image qualities and a high target/background ratio. Received 16 October and in revised form 17 December 1997     

Cell labelling with colloidal substances in whole blood

A method for the labelling of leucocytes with ^99mTc-colloid is described. The labelling can be done in samples of whole blood, because the colloid is only taken up by the phagocytosing cells, the monocytes and the granulocytes. The part of the colloid that is not phagocytosed is brought to a soluble state with Na-citrate, so that only the phagocytosed colloid is reinjected. The labelling efficiency with this method is between 80% and 90%. Measurements of the activity in the leucocytes 3 h after reinjection, have shown that at least 50% of the labelled cells are at this time still available in the blood pool.The clinical results on 32 patients with the tentative diagnosis of an abdominal abscess and on 42 patients with the tentative diagnosis of septic loosening of an endoprosthesis have shown that the labelled leucocytes are very well suited to show up local foci of inflammation.     

Rapid measurement of blood leakage during regional chemotherapy

In order to avoid complications after regional chemotherapy (isolated hyperthermic perfusion) of the extremities, rapid measurement of blood leakage from the extracorporeal to the systemic circulation is important. A method using technetium-99m in vivo red blood cell (RBC) labelling is reported that provides results within 3 min. Blood samples drawn from the systemic and the extracorporeal circulation were measured for ^99mTc activity using a mobile well counter, and the leakage values calculated. The mean result was 7.6%±6.5%/15 min (n=209). The corresponding flow rate was 100.2±85.7 ml/15 min (mean ± SD). The values for isolation perfusion of the upper and the lower extremities are compared. The leakage results using ^99mTc RBC labelling were correlated with other blood pool markers. Iodine-125 human serum albumin and indium-113 m transferrin were administered in subgroups of 4 and 19 patients simultaneously. Using linear regression, the coefficient of correlation was 0.72 for ^99mTc/^113mIn and 0.58 for ^99mTc/¹²⁵I. Comparison with the alternatives suggests that the rapid method of leakage measurement after ^99mTc RBC labelling can be considered one of the most practicable and reliable methods available.This paper is dedicated to Prof. E. Oberhausen, Homburg/Saar, on the occasion of his 65th birthday Correspondence to: C. Alexander