99Tcm and 111In leucocyte scintigraphy in inflammatory bowel disease.

A comparative study of 99Tcm-hexamethylpropyleneamine oxime (HMPAO) and 111In leucocyte scintigraphy was performed in inflammatory bowel disease. Two hundred and thirty-four patients were studied, 146 had 99Tcm-HMPAO, 82 had 111In and six had both. The sensitivity, specificity and accuracy of the 99Tcm leucocyte scan were 96, 97 and 97%, respectively, and 96, 97 and 97%, respectively, for the 111In leucocyte scan. 99Tcm-HMPAO leucocytes demonstrated similar diagnostic accuracy to 111In-labelled leucocytes with improved image quality and reduced radiation dose.     

Comparison of 99Tcm-HMPAO and 111In-oxine labelled granulocytes in man: first clinical results

The in vitro and in vivo behaviour of 99Tcm-HMPAO (hexamethylpropyleneamineoxime) (n = 12) and 111In-oxine (n = 11) labelled granulocytes, isolated by density-gradient centrifugation (Metrizamide/plasma gradients), was compared in patients with suspected inflammatory diseases. The in vitro elution of both labels and the viability of the labelled cells (99Tcm, 98.5%; 111In, 96.5%) was comparable but the labelling efficiency was different (99Tcm, 44 +/- 13%; 111In, 72.5 +/- 5.5%). In vivo, the lung (t1/2 max: 7.7 min), liver and spleen perfusion patterns were nearly identical; the image quality for detail in 99Tcm scans was superior to 111In images. The blood disappearance curves of 99Tcm and 111In were comparable. In the small number of patients examined all infections could be diagnosed correctly, without false-positive or false-negative results. Disadvantageous is the renal excretion of 99Tcm complexes (3+% over 20 h) with kidney and bladder activity from the beginning of the study. The biliary excretion in half of the patients (n = 6) with unspecific positive small and large bowel visualization and the late intestinal excretion also render the diagnosis more difficult. The recommended best imaging times for abdominal and retroperitoneal inflammations are 30 min to 2 h after injection. Late scans in septic prosthetic joints have disproportionate long acquisition times. As a potential cell labelling compound, 99Tcm-HMPAO has a promising future in comparison to 111In scans because of the good availability of 99Tcm, the image quality and the lower radiation exposure to the patient when lower activities for the early diagnosis of abdominal inflammatory diseases are reinjected.     

Osteomyelitis

The use of 111In-labelled granulocyte scintigraphy is recognized as a reliable method for detecting osteomyelitis and has similar sensitivity and significantly increased specificity compared to bone scintigraphy and 67Ga studies. Recent published work using pure granulocytes labelled with 111In tropolonate to detect osteomyelitis resulted in sensitivity of 100% and specificity of 92%. 99Tcm as an alternative granulocyte label offers advantages of convenience, lower radiation dose and higher image resolution. We have scanned 20 patients with suspected osteomyelitis using autologous granulocytes labelled with 99Tcm hexamethylpropyleneamineoxime (HMPAO), 12 of whom had prosthetic joints. The scan results were correlated with clinical, radiographic, microbiological and histological findings. Sensitivity was 100% and specificity was 93% which compares favourably with results obtained using 111In-labelled granulocytes. We believe that labelled granulocyte scintigraphy is a useful investigation in the diagnosis of osteomyelitis and that 99Tcm HMPAO appears to be at least as useful as 111In as the labelling agent.     

The development of a clinical protocol for the radiolabelling of mixed leucocytes with 99Tcm-hexamethylpropyleneamine oxime

To devise a protocol for the radiolabelling of mixed leucocytes with 99Tcm-hexamethylpropyleneamine oxime (99Tcm-HMPAO), the effect of HMPAO concentration, cell concentration, plasma concentration and anticoagulant on the uptake of the lipophilic complex was measured, together with the stability of the 99Tcm on the labelled cells. It was found that cell uptake was rapid, independent of HMPAO concentration over the range 20-80 micrograms ml-1, but dependent on cell and plasma concentration. Incubation of mixed leucocytes from 85 ml blood with 1 ml ACD-plasma and 4 ml 99Tcm-HMPAO containing 400 MBq 99Tcm for 10 min at room temperature gave optimum results and was used in 90 patients. The mean labelling efficiency, which was the amount of added 99Tcm incorporated by the cells, was 55% (+/- 13 S.D.), of which 77% were incorporated by the granulocytes, 17% by the mononuclear cells and 6% by the platelets and erythrocytes. During a 1 h incubation in plasma 9% (+/- 4% S.D.) of the 99Tcm were released from the labelled mixed leucocytes. This occurred predominantly from the mononuclear cells.     

Localization of inflammation with 111In IgG and 99Tcm albumin colloid labelled leukocytes in a rabbit model

Localization of inflammation with two recently described radiotracers, 111In-labelled polyclonal IgG and 99Tcm albumin colloid labelled leukocytes (Tc-WBC), was studied. Accumulation of activity was compared with 111In-labelled leukocytes (In-WBC) using 131I human serum albumin as a control. Ratios of activity in a chemically induced abscess in the thigh of rabbits compared with normal muscle tissue were measured. The results showed that all agents localize in inflammation but Tc-WBC consistently localizes to a greater degree than the other agents. At 2 h the inflammed-to-normal ratios for Tc-WBC were 4, IgG 2.1, albumin 1.9 and In-WBC 1.7. The pattern of the ratios remained similar over the 18 h period of the study. The short time in which leukocytes can be labelled and the quality of the images obtained suggest that Tc-WBC imaging is the method of choice for this model.     

Clinical comparison of 99Tcm-HMPAO labelled leucocytes and 99Tcm-nanocolloid in the detection of inflammation

Forty-five patients with various inflammatory diseases were imaged with 99Tcm-HMPAO labelled leucocytes and 99Tcm-nanocolloid within 7 days. The overall sensitivity of 99Tcm-leucocytes was 97% and that of 99Tcm-nanocolloid 59% and both agents had a 100% specificity. The 99Tcm-leucocyte method showed reliable results in various inflammatory and infectious conditions, and seems suitable as a primary imaging method. On the contrary, 99Tcm-nanocolloid cannot be recommended for use in inflammatory bowel diseases, soft tissue abscesses or prosthetic vascular graft infections. However, 99Tcm-nanocolloid gave reliable information in inflammatory and infectious bone and joint diseases in which it had a 90% sensitivity and 100% specificity. In those lesions the 99Tcm-nanocolloid method may be useful, because it is simple, fast and cheap. Yet, further evaluation is needed.     

A rapid method for the preparation of 99Tcm hexametazime-labelled leucocytes

99Tcm (+/-)-hexamethylpropyleneamineoxime (HMPAO) (99Tcm Hexametazime) has been recently reported as an alternative for labelling leucocytes. For reasons of convenience, radiation dose and image quality, many workers have welcomed this novel approach except for its complicated labelling protocol and venesection of 100 ml. This technique has been modified to give a simpler routine in-house labelling technique. It has three advantages: only about 20 ml of blood is required, the labelling time is just under 1 h and high yields of labelled leucocytes are obtained (mean of 500 MBq per injection dose). The properties of labelled leucocytes using this modified method are; 80% granulocyte-bound radioactivity, a rapid lung transit and a blood granulocyte recovery of 40% at 30 min similar to those described previously. The viability of the labelled leucocytes was tested and confirmed in vitro using a migration technique and in vivo by showing no lung retention on early imaging and high splenic uptake. A rapid in-process chromatography assessment procedure for regulating the protocol has been developed. Successful abscess imaging by 4 h has been achieved in 21 patients with normal results in another 22 patients without abscesses. This simpler method should encourage a more widespread application of scintigraphy using radiolabelled granulocytes.     

99Tcm-HMPAO leucocyte scintigraphy in the diagnosis of bone infection

The utility of 99Tcm-HMPAO leucocytes has been studied in combination with 99Tcm-MDP bone scanning in the diagnosis of bone infection in a series of 50 patients with a clinical suspicion of bone infection. Thirty-three patients were referred to our Service from the Department of Orthopaedic Surgery (Group A) and seventeen from the Infectious Disease Unit (Group B). A total of 52 lesion sites were studied. The leucocyte and bone studies were performed within four days. The leucocyte scan was obtained at 30-60 min and 4-6 h after i.v. injection of 370 +/- 74 MBq of 99Tcm-HMPAO leucocytes. After confirming the scintigraphic findings, the results obtained were: Group A, 12 true positive, 21 true negative and 2 false positive; and in Group B, 5 true positive, 9 true negative and 4 false negative. The overall sensitivity was 80.9% with a specificity of 93.7%. Although the high bone marrow activity seen with 99Tcm-HMPAO leucocytes may reduce sensitivity, very good results were obtained in bone infection. The use of 99Tcm means great progress in the radiolabelling of white blood cells in terms of availability and better image quality. The combination of 99Tcm-HMPAO leucocytes and 99Tcm-MDP can be recommended as one of the most suitable methods for use in the diagnosis of bone infection, especially in patients with previous bone disease.     

Imaging of inflammation with granulocytes labelled in vivo

We have looked for inflammatory lesions in 37 patients using granulocytes labelled in vivo. For the purpose of cell labelling we used a monoclonal murine antibody reacting with NCA and carcinoembryonic antigen (CEA) (BW 250/183) labelled with 99Tcm. All abscesses and other inflammatory lesions were visualized with excellent quality scintigrams between 2 and 6 h after the injection. As the antibody can be stored in a freeze-dried form and labelled at any time in any department of nuclear medicine with 99Tcm without cell separation being necessary, the method appears to be suitable even for use in acute diagnosis. No side effects have so far been found.     

Radiolabelled granulocytes in inflammatory bowel disease: diagnostic possibilities and clinical indications

Radiolabelled granulocytes in chronic inflammatory bowel diseases (CIBD) are able to diagnose the disease extent and assess the disease activity. It may be performed with 111In oxine- and 99Tcm hexamethylpropyleneamineoxime (HMPAO)-labelled cells. The granulocyte scan localizes inflamed bowel segments with an accuracy comparable with radiology and endoscopy including biopsy of the bowel (r = 0.95; P less than 0.001). The specificity of the scan for diseased segments is near 100%, the sensitivity 92%. A three-phase white blood cell scan (imaging: 0.5, 4, 20 h post injection) allows differentiation of diseased bowel segments from abscesses and fistulas. False positive results are possible in necrotic carcinomas. The 99Tcm HMPAO scan shows rapid renal and delayed biliary and intestinal excretion of tracer. In this way diagnostic problems arise in the small pelvis. Because of the intestinal and biliary excretion, early images should be obtained (0.5-2 h post injection). Later scans with 99Tcm HMPAO are of minor importance. The disease activity can very specifically be assessed by the determination of the percentage faecal 111In excretion (96 h faecal collection). Active and non-active diseases can be clearly differentiated. We found no correlation with the subjectively influenced Crohn's disease activity index (CDAI) (r = 0.25; P greater than 0.05), but good correlations with the Dutch Index (van Hees: r = 0.67), ESR (r = 0.69), serum albumin (r = -0.54) and orosomucoid (r = 0.65). The percentage faecal excretion correlates well with the histologically estimated leucocytic bowel infiltration. Because of the intestinal 99Tcm HMPAO excretion the determination of faecal excretion is pointless in 99Tcm studies.(ABSTRACT TRUNCATED AT 250 WORDS)     

99Tcm hexamethylpropyleneamineoxime (HMPAO) as a platelet label: evaluation of labelling parameters and first in vivo results

The lipophilic 99Tcm hexamethylpropyleneamineoxime (HMPAO) complex can be used for labelling platelets as well as granulocytes. For the evaluation of optimal labelling parameters, platelets were isolated according to standard isolation procedures. The labelling efficiency (%) depends on incubation temperature (22 degrees C: 40%; 37 degrees C: 50%), incubation time (3 min: 20%; 25 min: 55%) and the incubation medium (plasma: 40%; saline: 50%). The 60 min 99Tcm elution from the platelets is around 8%. The platelet recovery, used as a quality parameter, is around 25 +/- 4% and is stable for at least 240 min. The high elution rate from the platelets leads to renal excretion of the label and so to significant kidney and bladder activity. Intestinal excretion of the label can also be frequently demonstrated. Fresh thrombotic lesions can usually be detected 4 h after reinjection of the labelled platelets, and in some patients as early as 1 h after reinjection of the platelets. In conclusion, 99Tcm 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 99Tcm.     

99mTc-human immunoglobulin (HIG)--first results of a new agent for the localization of infection and inflammation

Technetium (99mTc) labelled, polyclonal human immunoglobulin (HIG) is a new agent that detects focal infection and inflammation. This new agent was compared in 40 patients with the accepted standard, namely 111In-oxine-labelled leucocytes. This comparison resulted in a sensitivity of 94% and a specificity of 96% for 99mTc-HIG when 111In-oxine leucocytes were defined as giving the true result. The new agent was shown to localize both sepsis and active inflammatory bowel disease (IBD). There was 100% concordance in the 16 patients with IBD who were imaged with both 99mTc-HIG and 111In-oxine leucocytes. Discordant results were obtained in one case of suspected osteomyelitis, which was false-positive on the 99mTc-HIG scan, and one case of pyrexia of unknown origin when the 99mTc-HIG was false-negative and the 111In-oxine leucocyte scan demonstrated accumulation of tracer in the caecum at 24 h post-injection. Normal distribution for 99mTc-HIG demonstrated activity in the kidneys and bladder and that 50% of the tracer is cleared through the kidneys during the first 24 h post-injection. There were no major or minor side-effects.     

Indium-111 labelled lymphocytes: isotope distribution and cell division

Since lymphocytes continue to proliferate and divide in vivo, it is important to determine the fate of a radionulide following lymphocyte labelling. Using the mixed lymphocyte reaction (MLR), we induced indium-111 labelled lymphocytes from a specific in-bred rat strain (AS) to divide and then observed the subsequent¹¹¹In distribution between cells and supernatant. L10 and L12.4 cells, which are allospecific CD4+ T lymphocytes from the AS rat, were stimulated in the MLR by antigen-presenting cells from the August rat, a different strain. We labelled L10 or L12.4 lymphocytes on day 0, the first day of the stimulation cycle, and continued to culture the lymphocytes in vitro. The proliferation of the cells was estimated according to their increase in number. The distribution of¹¹¹In between cell and supernatant fractions and between viable and dead (but intact) cells was measured in the cell suspension each day after labelling. The metabolic activity of¹¹¹In-labelled lymphocytes was compared with control cells by measuring their uptake of fluorine-18 fluorodeoxyglucose ([¹⁸F]FDG).¹¹¹In-labelled lymphocytes showed a poor proliferative response compared with control cells 24–48 h after labelling but increased in number after this time. From 24 to 72 h, about 70% of¹¹¹In was in the supernatant but only about 5%–10% was associated with intact dead cells. These dead cells tended to retain their¹¹¹In, losing less than 30% per day, suggesting that¹¹¹In in the supernatant was the result of active elimination from viable cells. Moreover, 24 h after culture, considerably more¹¹¹In was associated with viable than with dead lymphocytes, although over the next few days this distribution reversed.¹¹¹In-labelled lymphocytes took up more [¹⁸F]FDG than control cells at 24 h but not at 0 or 72–96 h; the maximum [¹⁸F]FDG uptake coincided with the greatest reduction in cell number. Furthermore, [¹⁸F]FDG uptake correlated with the initial¹¹¹In burden in lymphocytes labelled with¹¹¹In 24 h previously. The results are consistent with active elimination of¹¹¹In by¹¹¹In-labelled lymphocytes. The energy requirements for this are diverted away from cell division, thereby increasing the probability of cell death. As lymphocytes become¹¹¹In deplete, they recover their capacity to proliferate and their risk of death decreases. These findings have important implications for¹¹¹In-labelled lymphocyte scintigraphy, suggesting that cells remaining viable immediately after labelling will either subsequently die or alternatively eliminate the label.     

Radiochemistry and biostability of autologous leucocytes labelled with 99mTc-stannous colloid in whole blood

Autologous leucocytes were labelled in whole blood by phagocytic uptake of 99mTc-stannous colloid. This colloid has a mean particle size of 1.5 micron and labels leucocytes with 81% efficiency. Individual cell uptakes were: granulocytes 42%, monocytes 39%. Isotonic sodium citrate added after the labelling procedure did solubilise excess colloid but was not necessary for adequate removal of excess colloid. Labelled leucocytes were shown in vitro to be viable and maintain normal bactericidal and chemotactic capacity. Biodistribution, clearance rates and dosimetry are presented. These results indicate that autologous leucocytes can be efficiently labelled with 99mTc-stannous colloid with good residual cell function.     

White cells radiolabelled with 111In and 99Tcm--a study of relative sensitivity and in vivo viability

In this study a comparison between the classical (111In oxine) and the newer (99Tcm HMPAO) technique of labelling leucocytes is reported. The behaviour in vivo and the relative sensitivity in the detection of infection (chest and bone) and inflammatory bowel disease (IBD) is presented. Simultaneous dual-radionuclide gamma camera acquisition methodology was applied to study 99 patients, 18 with chest infection, 26 with bone infection, 41 with IBD and 14 with other pathological conditions. The mean (1 SD) 50% washout time from the lungs was 483.03 (79.10) s for 99Tcm HMPAO-labelled white blood cells and 475.85 (83.79) s for 111In oxine-labelled white cells (r = 0.81). Concordance between the two techniques was 94% in the chest-infection group of patients, 88% in the bone-infection group and 71% in the localization of IBD.     

Simultaneous administration of111In-human immunoglobulin and99mTc-HMPAO labelled leucocytes in inflammatory bowel disease

Technetium-99m hexamethylpropylene amine oxime (HMPAO) labelled leucocytes and indium-111 polyclonal immunoglobulin (IgG) were simultaneously injected into a group of 27 patients routinely referred for the investigation of inflammatory bowel disease (IBD). Ten-minute anterior abdomen and tail on detector views were obtained at 30 min, 4 h and 24 h p.i. of both tracers. The diagnosis of IBD was obtained in all cases by endoscopy with biopsy and/or surgery. Images were blindly evaluated by two experienced observers who only knew of the clinical suspicion of IBD. IBD was confirmed in 20 patients (12 with Crohn's disease and eight with ulcerative colitis). Sensitivity, specificity and accuracy were 100%, 85% and 96% respectively for labelled leucocytes and 70%, 85% and 74% for IgG. Both IgG and leucocyte scans were normal in six out of seven patients in whom a diagnosis of IBD was excluded; the remaining patient, with ischaemic colitis, was falsely positive with both agents. As far as disease extension is concerned, the IgG study localized 27 diseased segments, whereas 49 were seen with the leucocyte study. Eighty-four segments were normal and 25 showed tracer uptake with both agents. Twenty-four were positive only with the leucocyte study and two were positive only with the IgG study. Agreement between the agents was 80.7%. These results confirm that¹¹¹In-human polyclonal scintigraphy is less sensitive than^99mTc-HMPAO scintigraphy both for the diagnosis of IBD and in the evaluation of disease extension. Nevertheless, if leucocyte labelling is not available, labelled IgG can be used only for diagnostic purposes.     

Evaluation of white cell scintigraphy using indium-111 and technetium-99m labelled leucocytes

Indium-111 oxine labelled leucocyte (¹¹¹In oxine leucocyte) scintigraphy is the test of choice in detecting occult infection and localising focal inflammation. ¹¹¹In oxine labelling is technically difficult and expensive and leucocyte labelling with technetium-99m stannous colloid (^99mTc Sn colloid) has been considered to be an alternative. Leucocytes from 40 cases referred for investigation of occult infection or localisation of inflammation were simultaneously labelled with ¹¹¹In oxine and ^99mTc Sn colloid with dual isotope acquisition performed at 1, 3 and 24 h. Twenty-four hour ^99mTc Sn colloid scans were corrected for ¹¹¹In downscatter. Each case was independently interpreted by two experienced observers. Twentyone patients demonstrated positive ¹¹¹In oxine leucocyte scans. Using ¹¹¹In oxine leucocyte scans as the gold standard, ^99mTc Sn colloid leucocyte scanning had an overall sensitivity of 86% and a specificity of 95%. Clinical follow-up verified that three patients had false negative ^99mTc Sn colloid leucocyte scans and one patient had a false positive. Further clinical evaluation of ^99mTc Sn colloid labelled leucocytes is required before they can become a reliable replacement for ¹¹¹In oxine leucocytes. Correspondence to: S. Boyd     

Radiochemistry and biostability of autologous leucocytes labelled with 99mTc-stannous colloid in whole blood

Autologous leucocytes were labelled in whole blood by phagocytic uptake of ^99mTc-stannous colloid. This colloid has a mean particle size of 1.5 m and labels leucocytes with 81% efficiency. Individual cell uptakes were: granulocytes 42%, monocytes 39%. Isotonic sodium citrate added after the labelling procedure did solubilise excess colloid but was not necessary for adequate removal of excess colloid. Labelled leucocytes were shown in vitro to be viable and maintain normal bactericidal and chemotactic capacity. Biodistribution, clearance rates and dosimetry are presented. These results indicate that autologous leucocytes can be efficiently labelled with ^99mTc-stannous colloid with good residual cell function.     

Comparison of 99Tcm complexes (NEP-DADT, ME-NEP-DADT and HMPAO) with 123IAMP for brain SPECT imaging in dogs

Several new lipophilic 99Tcm complexes have recently been described as alternatives to N-isopropyl (123I) iodoamphetamine (123IAMP) for measurement of regional cerebral blood flow (RCBF). In this study we have compared brain uptake and blood clearance of 99Tcm-N-ethylpiperidine-diamino dithiol (99Tcm-NEP DADT), its 4-methylated derivative (99Tcm-Me-NEP-DADT) and 99Tcm-hexamethyl-propylene-amine-oxime (99Tcm-HMPAO) with that of 123IAMP in two dogs. Single photon emission tomography (SPECT) was employed to measure brain accumulation and retention of the four radiopharmaceuticals. Cerebral uptake of the 99Tcm complexes (0.8-1.1%) was lower than that of 123IAMP (1.6% of the injected dose). There was considerable extracerebral activity in the dog's head, especially in the olfactory and snout regions. Because of slow blood clearance 99Tcm-HMPAO showed high uptake in these regions. Brain uptake of 99Tcm-HMPAO reached a plateau 5 to 10 min after intravenous injection and remained constant for the entire study period (1 h). 99Tcm-NEP-DADT, on the other hand, showed significant clearance from the brain after reaching maximal uptake at 10 to 15 min after injection. However, brain imaging with these agents was possible during the first 20 min. The mechanism of brain uptake, as well as the relationship between brain uptake and RCBF need to be evaluated for each of the four radiopharmaceuticals.     

99Tcm-labelled HSA-nanocolloid versus 111In oxine-labelled granulocytes in detecting skeletal septic process

Fifty-seven investigations of the skeletal system were performed on 54 patients, using a 99Tcm-labelled nanometer-sized HSA colloid in a crossover comparison with 111In oxine-labelled granulocytes for the detection of sites of infection. The findings were in agreement in 55 out of 57 investigations (96.5%). Based on 44 studies in which a final clinical diagnosis was obtained, both methods were found to display the same specificity (93%), whilst the sensitivity of 99Tcm nanocolloid scintigraphy (87%) was slightly higher than that obtained with 111In leucocyte scintigraphy (81%). In our opinion, 99Tcm nanocolloid is easier to use and the total duration of the investigation is considerably shorter. The use of 99Tcm is scintigraphically more advantageous and, with the dosage required, the absorbed radiation dose to the red bone marrow is three times lower than with 111In granulocytes. For the detection and therapy monitoring of osteomyelitis, as well as for the investigation of arthroplasties suspected of infective loosening, we consider scintigraphy with 99Tcm nanocolloid to be equivalent to leucocyte scintigraphy. Identical findings were obtained with both tracers in suspected spondylodiscitis.