Benzalkonium runoff from roofs treated with biocide products - In situ pilot-scale study.

Roof maintenance practices often involve the application of biocide products to fight against moss, lichens and algae. The main component of these products is benzalkonium chloride, a mixture of alkyl benzyl dimethyl ammonium chlorides with mainly C12 and C14 alkyl chain lengths, which is toxic for the aquatic environment. This paper describes, on the basis of an in-situ pilot scale study, the evolution of roof runoff contamination over a one year period following the biocide treatment of roof frames. Results show a major contamination of roof runoff immediately after treatment (from 5 to 30 mg/L), followed by an exponential decrease. 175-375 mm of cumulated rainfall is needed before the runoff concentrations become less than EC50 values for fish (280 µg/l). The residual concentration in the runoff water remains above 4 µg/L even after 640 mm of rainfall. The level of benzalkonium ions leaching depends on the roofing material, with lower concentrations and total mass leached from ceramic tiles than from concrete tiles, and on the state of the tile (new or worn out). Mass balance calculations indicate that a large part of the mass of benzalkonium compounds applied to the tiles is lost, probably due to biodegradation processes.

Disease- and treatment-related elevation of the neurodegenerative marker tau in children with hematological malignancies.

Children acquire neuropsychologic dysfunctions after chemotherapy for hematologic malignancy. In this study, putative changes in levels of CSF-tau (a marker of neural dysintegrity) in leukemic children prior to and during chemotherapy were studied. Cerebrospinal fluid (CSF) samples were obtained before and during treatment from patients with B cell non-Hodgkin's lymphoma (NHL, n = 10), non-B cell acute lymphoblastic leukemia/NHL (non-B-ALL, n = 48), acute myeloid leukemia (AML, n = 9), other malignant diseases (n = 9), and six control children. A sandwich-type ELISA (INNOTEST hTAU-Ag) was used for measuring CSF-tau. Sixteen out of 50 patients with hematological malignancies, including the patients with proven leukemic CNS invasion, already showed high CSF-tau levels at baseline (>300 pg/ml). The pre-induction treatment for non-B-ALL, consisting of only corticosteroids and methotrexate (MTX), resulted in a significant increase of tau at day 8 (on average to 535 pg/ml). Larger increases as compared to baseline levels of CSF-tau were observed in patients treated for B-NHL with systemic vincristine, corticosteroids and cyclophosphamide, and intrathecal MTX (mean 776 pg/ml at day 8). In two AML patients with CNS invasion, CSF-tau increased during chemotherapy up to 1,500 and 948 pg/ml, respectively. In one non-B-ALL patient with MTX-induced clinical neurotoxicity, CSF-tau was above the detection limit of 2,000 pg/ml. Almost one-third of the patients with hematological malignancies had elevated CSF-tau levels at diagnosis. Transient high levels of CSF-tau, reaching levels observed in other neurodegenerative disorders, were observed during induction chemotherapy for non-B-ALL, B-NHL and CNS+ AML. The clinical implications of both observations will be the subject of further study.

In vitro and in vivo biocompatibility of dextran dialdehyde cross-linked gelatin hydrogel films.

The biosafety of a new hydrogel wound dressing material consisting of dextran dialdehyde cross-linked gelatin was evaluated (i) in vitro in cultures of dermal fibroblasts, epidermal keratinocytes, and endothelial cells, three cell types which play a major role in the process of cutaneous wound healing, and (ii) in vivo by subcutaneous implantation studies in mice. The cytotoxicities of this hydrogel, two semi-occlusive polyurethane dressings (Tegaderm and OpSite), and a hydrocolloid dressing (DuoDERM) were compared by measuring cell survival with the tetrazolium salt reduction (MTT) assay after incubations of the wound dressing samples for up to 6 d, in the presence of--but not in direct contact with--the cells. In vitro, the degree of cytotoxicity of the new hydrogel was greater in keratinocyte cultures than in fibroblast and endothelial cell cultures, and increased upon longer incubation time. In keratinocyte cultures, the semi-occlusive polyurethane dressings, the hydrocolloid, and the hydrogel dressings induced low, high and acceptable degrees of cytotoxicity, respectively. The toxicity of the isolated hydrogel components was assessed in Balb MK keratinocyte cultures. In these cells, epidermal growth-factor-stimulated thymidine incorporation into DNA was higher in the presence of gelatin. By contrast, concentrations of dextran dialdehyde as low as 0.002% were found to significantly decrease thymidine incorporation (P < 0.01). Subcutaneous implantation studies in mice showed that in vivo the hydrogel was biocompatible since the foreign body reaction seen around the implanted hydrogel samples was moderate and became minimal upon increasing implantation time. These results indicate that dextran dialdehyde cross-linked gelatin hydrogels have an appropriate biocompatibility.

In vitro release characteristics of bioactive molecules from dextran dialdehyde cross-linked gelatin hydrogel films.

Hydrogel films, prepared by cross-linking of gelatin with dextran dialdehydes (weight ratio 2:1), and containing either fluorescein isothiocyanate dextran (Mw 70000) or polypeptides were evaluated in terms of their release characteristics and mechanical properties upon increasing storage time at 4 degrees C. Important changes in release kinetics and mechanical properties of the cross-linked gelatin films were observed, especially during the first week after the hydrogel production. Rheological and NMR measurements showed that the mechanical properties of the gelatin hydrogel films were improved with increasing storage time. It appeared that the process of chemical cross-linking and physical structuring of the gelatin hydrogel matrix did not occur instantaneously and substantially influenced the polypeptide release patterns. Cross-linked gelatin hydrogels were found to be appropriate release systems for medium-term sustained delivery of biologically active epidermal growth factor (EGF), but release characteristics were strongly dependent on the nature of the protein which was incorporated.

Cerebrospinal fluid tau protein as a biochemical marker for Alzheimer's disease: a community based follow up study.

OBJECTIVES: Biochemical markers for Alzheimer's disease would be of great value, especially to help in diagnosis early in the course of the disease. A pronounced increase in CSF tau protein (CSF-tau) is found in most patients with Alzheimer's disease. However, the specificity has to be further studied, as an increase in CSF-tau has also been found in other dementias, especially in vascular dementia. As most previous CSF studies have been based on selected inpatients, it was considered of special interest to examine the diagnostic potential of CSF-tau in a community population based sample of consecutive patients with dementia. Such patient material has been examined at the Piteå River Valley Hospital in Northern Sweden since 1986, and includes all those with memory disturbances in the community. The aim was also to study if an increase in CSF-tau is found early in the disease process, and whether CSF-tau changes during the progression of disease. PARTICIPANTS: Community population based sample of 75 demented patients (43 with Alzheimer's disease, 21 with vascular dementia, and 11 with mixed Alzheimer's disease/vascular dementia), 18 healthy subjects, and 18 neurological controls. A follow up investigation (including analysis of a new CSF sample) was performed in all patients after about one year. MAIN OUTCOME MEASURES: Concentrations of total (both normal tau and PHF-tau) tau in CSF, clinical measures (duration and severity of dementia), and apoE polymorphism. RESULTS: CSF-tau was markedly increased in Alzheimer's disease, 41/43 (95%) patients had values above the cut off level (mean+2 SD) in controls (306 pg/ml). High CSF-tau concentrations were also found in most patients with vascular dementia, preferentially in patients with vascular dementia without progressive leukoaraiosis on CT, whereas patients with vascular dementia with progressive leukoaraiosis had normal CSF-tau. Concentrations of CSF-tau were stable at one year follow up in both patients with Alzheimer's disease and patients with vascular dementia, and there was no correlation between CSF-tau and either duration or severity of dementia. CONCLUSIONS: The findings confirm the high sensitivity of CSF-tau for the diagnosis of Alzheimer's disease, but high CSF-tau was also found in vascular dementia, resulting in a lower specificity. However, high CSF-tau is preferentially found in patients with vascular dementia without progressive leukoaraiosis, which may constitute a group with concomitant Alzheimer's disease pathology. High CSF-tau may be present during the whole course of the disease in Alzheimer's disease. Possibly, therefore, the same high CSF-tau concentrations may be present before the onset of clinical dementia. Follow up studies on such patients will tell whether analysis of CSF-tau is useful as a biochemical marker for early Alzheimer's disease.

Generation of a mouse tumor necrosis factor mutant with antiperitonitis and desensitization activities comparable to those of the wild type but with reduced systemic toxicity.

In this study, we investigated whether the recently identified lectin-like domain of tumor necrosis factor (TNF) is implicated in its biological activities on mammalian cells. To this end, a mouse TNF (mTNF) triple mutant, T104A-E106A-E109A mTNF (referred to hereafter as triple mTNF), lacking the lectin-like affinity of mTNF for specific oligosaccharides, was compared with the wild-type molecule for various TNF effects in vitro and in vivo. The triple mTNF displayed a 50-fold-reduced TNF receptor 2 (TNFR2)-mediated bioactivity but only a 5-fold-reduced TNFR1-mediated bioactivity in vitro. The specific activity of the triple mutant on L929 fibrosarcoma cells was slightly reduced compared with that of the wild type. We subsequently assessed the systemic toxicity of triple versus wild-type mTNF, since TNFR2 is partially implicated in this activity. The triple mTNF had a significantly reduced toxicity compared with that of wild-type mTNF in vivo. Moreover, we compared the effects of the triple and the wild-type mTNFs in TNFR1-mediated phenomena, such as (i) induction of tolerance towards a lethal mTNF dose and (ii) protective activity in cecal ligation and puncture-induced septic peritonitis. No significant differences between the mutant and wild-type forms were observed. In conclusion, these results indicate that triple mTNF, lacking TNF's lectin-like binding capacity, has reduced systemic toxicity but retains the tolerance-inducing and peritonitis-protective activities of wild-type mTNF.

Transmembrane segments critical for potassium channel function.

The relative contribution of the transmembrane segments in the alpha-subunit of Shaker-type potassium channels was investigated in relation to potassium channel function. Starting from a wild-type Kv1.1 channel, four different deletion mutants were made, missing respectively transmembrane segments S1 and S2, S2 and S3, S1 to S3, and S1 to S4. To ensure the assembly of the different subunits, the hydrophylic N-terminal domain was always conserved. The lack of transmembrane segments S1 to S4 converts a depolarization-activated WT Kv1.1 channel with outward rectification into a hyperpolarization-activated channel with inward rectification. In contrast, mutant channels missing transmembrane segments S1 and S2, S2 and S3, or S1 to S3 did not reveal functional expression.

The phosphorylation state of tau in the developing rat brain is regulated by phosphoprotein phosphatases.

The paired helical filaments (PHFs) in Alzheimer's disease neurofibrillary tangles are composed of PHF-tau which is thought to be hyperphosphorylated because several residues in postmortem samples of PHF-tau and human fetal tau are phosphorylated while the corresponding sites are not phosphorylated in autopsy-derived normal adult human brain tau. To determine how the phosphorylation of these sites is regulated, we isolated tau from rat brains at different embryonic and postnatal ages in the presence of okadaic acid to obtain tau in its most native in situ phosphorylation state. Fetal tau was highly phosphorylated from embryonic day 18 (E18) until postnatal day 11 (P11). Thereafter, the levels of fetal tau diminished as did its phosphorylation state concomitant with the appearance of the five adult tau isoforms. Several phosphorylation-dependent antibodies (i.e. AT270, AT8, AT180, T3P, and PHF1) that recognize PHF-tau also recognized these tau isoforms, albeit at reduced levels in the mature rat brain. This suggests that Thr172, Ser193, Thr222, Ser387, and Ser395 are normal sites of phosphorylation in rat brain tau. The inclusion of OK in the microtubule assembly buffers did not alter the ability of tau to bind microtubules at any age. However, phosphatases were activated and kinases were down-regulated in the rat brain after P12 since adult tau proteins were partially dephosphorylated at and beyond this time in the absence of OK. Protein phosphatase 2A (PP2A) and 2B (PP2B) activities in the adult rat brain extracts dephosphorylated tau efficiently, but protein phosphatases in extracts of the P6 rat brain did not have a similar effect. This suggests that the sensitivity of tau to OK after P12 may be regulated by the de novo induction of adult brain phosphatases. Finally, PP2A and/or PP2B in adult rat brain extracts dephosphorylated tau in a site-specific manner. Thus, PP2A and PP2B (or closely related phosphatases) may regulate the phosphorylation state of adult tau isoforms in vivo, and the generation of PHF-tau in the AD brain may result from the abnormal inactivation of similar phosphatases.

Characterization of a rat C6 glioma-secreted follistatin-related protein (FRP). Cloning and sequence of the human homologue.

A protein was isolated from rat C6 glioma-conditioned medium and was biochemically characterized. The heparin-binding protein has a native molecular mass of 55-75,000 Da, a molecular mass of 40-48,000 Da under denaturing conditions, and a pI of 5.0-6.0. Based on the determined partial amino acid sequences, the full lenght cDNA encoding the rat and human proteins were cloned. The cDNA sequences identified the isolated rat and human protein as the homologue of a recently reported mouse osteoblast-transforming-growth-factor-beta 1-inducible protein, encoded by the TSC-36 gene [Shibanuma, M., Mashimo, J., Mita, A., Kuroki, T. & Nose, K. (1993) Eur. J. Biochem. 217, 13-19]. Analysis of the human, rat and mouse amino acid sequences indicates that these proteins are highly conserved (> 92% sequence identity). Sequence similarities with follistatin and the follistatin-like domain of agrin are revealed. The relationship with follistatin and agrin points to possible common functions for the cloned follistatin-related proteins (FRP). The protein has no effect on the inhibitory action of transforming growth factor-beta 1, on CCl-64 cell growth.

Secretion of steroids, growth factors, and cytokines by immortalized mouse granulosa cell lines.

The expression and function of gonadotropin receptors, and the secretion of steroids, transferrin, and cytokines were investigated in three immortalized (single transfection with v-myc) mouse granulosa cell lines (GRM01, GRM01L, and GRM02). A dose-dependent increase in progesterone production was obtained in GRM01 and GRM02 cells after addition of LH, FSH, modulators of the adenylate cyclase enzyme system, and cAMP analogues. The LH-induced release of progesterone was already detectable in GRM02 cells after 8 h and was related to incubation time and cell number. Both epidermal growth factor (EGF) and transforming growth factor alpha (TGF alpha) induced the secretion of progesterone in GRM02 cells, while no effect was obtained with TGF beta. LH receptor concentration was highest in the GRM02 cell line. FSH receptor mRNA was visualized in GRM01 and GRM02 cells. Aromatase activity in GRM02 cells was induced by androgens and inhibited by aromatase inhibitors. Whereas all cell lines were able to secrete transferrin, only in GRM01 cells was transferrin secretion increased significantly by LH. FSH did not affect transferrin secretion in the three cell lines, in contrast to forskolin or 8-bromo-cAMP. The immortalized mouse granulosa cell lines were able to express and release several growth factors. The expression and secretion of activin, inhibin, TGF beta, EGF, TGF alpha, insulin-like growth factor II, fibroblast growth factor (acidic and basic), platelet-derived growth factor, and interleukin-6 suggest an autocrine or paracrine role for these factors in follicular differentiation and function. In conclusion, these cells, derived from mural granulosa cells and immortalized in a preovulatory state, can be used to study granulosa cell physiology or to study the role of granulosa cells and their derivatives in the process of follicular maturation, fertilization, and early embryonic development.

Characterization of immortalized mouse granulosa cell lines.

Cell cultures of primary mouse granulosa cells were transfected with a v-myc-containing plasmid, and the resulting stable cell lines were tested for their steroidogenic properties and physiologic status. Granulosa cells were obtained from 22-day-old NMRI mice injected with 8 IU pregnant mare serum gonadotropin i.p. 2 days earlier. In Passage 1 the cells were transfected with pSVv-myc using calcium phosphate precipitation or lipofectin. The 3 beta- and 17 beta-hydroxy steroid dehydrogenase activity was visualized in control cultures. The three cell lines obtained have been in culture for over 1 yr and have been subcultured for more than 90 passages. The cell line GRM01, with a doubling time of 37 +/- 3 h and a diploid modal chromosome number, produced progesterone, estradiol, as well as inhibinlike and activinlike material under basal conditions. A combination of follicle-stimulating hormone and luteinizing hormone was able to increase the secretion of progesterone. GRM01L, a fast growing clone of the GRM01 line with a doubling time of 10 +/- 1 h, retained only the capacity to produce activinlike material and transforming growth factor-beta, and it was the only one with a tumorigenic capacity. Epidermal growth factor, insulin, and interleukin-6 were able to induce the [3H]thymidine incorporation into DNA in these two cell lines. GRM02, with a doubling time of 36 +/- 2 h and a hypertriploid modal chromosome number, produced progesterone and activinlike and inhibinlike material. Follicle-stimulating hormone and luteinizing hormone were able to enhance the secretion of progesterone. For this cell line, only insulin was shown to induce [3H]thymidine incorporation into DNA.

Detection of tau proteins in normal and Alzheimer's disease cerebrospinal fluid with a sensitive sandwich enzyme-linked immunosorbent assay.

Alzheimer's disease is a progressive degenerative dementia characterized by the abundant presence of neurofibrillary tangles in neurons. This study was designed to test whether the microtubule-associated protein tau, a major component of neurofibrillary tangles, could be detected in CSF. Additionally, we investigated whether CSF tau levels were abnormal in Alzheimer's disease as compared with a large group of control patients. We developed a sensitive sandwich enzyme-linked immunosorbent assay using AT120, a monoclonal antibody directed to human tau, as a capturing antibody. With this technique, the detection limit for tau was less than 5 pg/ml of CSF. Using AT8, which recognizes abnormally phosphorylated serines 199-202 in tau, the detection limit was below 20 pg/ml of CSF. However, with AT8, we found no immunoreactivity in CSF, suggesting that only a small fraction of CSF tau contains the abnormally phosphorylated AT8 epitope. Our results indicate that CSF tau levels are significantly increased in Alzheimer's disease. Also, CSF tau levels in a large group of patients with a diversity of neurological diseases showed overlap with CSF tau levels in Alzheimer's disease.

Identification by microsequencing of lipopolysaccharide-induced proteins secreted by mouse macrophages.

The conditioned medium of the murine macrophage PU5.1.8 was analyzed by two-dimensional gel electrophoresis in order to detect LPS-induced proteins. Spots of interest were identified by microsequencing of internal peptides generated by limited in situ acid hydrolysis. In total conditioned medium, several monokines (TNF-alpha and macrophage inflammatory protein-1 alpha and 1 beta) were identified as LPS-induced spots. Because minor spots could be masked by the complexity of the 2-D pattern, conditioned medium was successively fractionated by zinc precipitation and affinity chromatography (Procion red and Con A agarose). Zinc supernatant fraction, Procion red flow-through, and Con A eluate fractions were further analyzed by 2-D gel electrophoresis for the presence of LPS-induced spots. In these fractions serum amyloid A3, lipocalin 24p3, cathepsin B, and plasminogen activator inhibitor-I were characterized as LPS-induced proteins secreted by macrophages. Lipocalin 24p3 protein was retrieved for the first time. In addition to these proteins that follow a classical secretory pathway, several cellular proteins (mainly ribosomal proteins) were retrieved as LPS-induced proteins in the conditioned medium. Control experiments argue against the obvious explanation that the latter observation is caused solely by cellular leakage.

Magnetic beads in suspension enable a rapid and sensitive immunodetection of human placental alkaline phosphatase.

The kinetics and efficiency of the interaction between placental alkaline phosphatase and a monoclonal antibody (laboratory number 327) were determined by immunoassay using microtitre plates or magnetic beads. While only up to 45% of placental alkaline phosphatase was bound to microwells precoated with this antibody, even after prolonged incubation, no less than 60% and 100% binding were reached using magnetic beads after 1 and 3 h incubations, respectively. High-molecular-mass placental alkaline phosphatase and complexed placental alkaline phosphatase forms were also completely bound to magnetic beads in the presence of deoxycholate (up to 9 g/l for serum samples). The assay sensitivity was improved up to 4-fold. In addition, 100% binding of the antigen was achieved during simultaneous incubation of magnetic beads, monoclonal antibody (125 micrograms/l), and placental alkaline phosphatase. This one-step enzymatic assay, based on magnetic beads, is an attractive alternative to the classic assay performed in microtitre plates, enabling rapid, precise, and sensitive antigen detection, and only necessitating a minimum of laboratory equipment.

Production of functionally active murine and murine::human chimeric F(ab')2 fragments in COS-1 cells.

We report here the synthesis, in nonlymphoid cells, of two functionally active recombinant F(ab')2 fragments directed against the tumor marker, human placental alkaline phosphatase (hPLAP). The truncated heavy chain (HC) sequences, E6Hf2 and E6Hy3f2, of the murine F(ab')2 fragment, E6F2, and of the murine::human chimeric F(ab')2 fragment, E6(Hy3,kappa)F2, respectively, were engineered by introducing an in-phase stop codon within the second constant domain of the corresponding parental HC sequence. The antibody-encoding genes were placed under control of the simian virus 40 late promoter and each HC sequence, together with the light chain (LC) sequence, was transiently expressed in COS-1 cells. The truncated HCs were correctly synthesized, processed and assembled with the murine LC and subsequently secreted into the culture medium as functionally active entities with stable hinge region interactions. These results indicate that, under the conditions used, the hinge region was sufficient for the formation of divalent molecules. However, Western blotting revealed the presence of hPLAP-binding half-molecules of E6F2, which was not the case for E6(Hy3,kappa)F2. Since E6F2 and E6(Hy3,kappa)F2 mainly differ by the length of their hinge region (22 and 62 aa residues, respectively) and the number of inter-HC disulfide bridges (four and eleven, respectively), it may be concluded that F(ab')2 fragments with an extended hinge region and several inter-HC disulfide bridges are formed more efficiently.

Affinity purification of human tau proteins and the construction of a sensitive sandwich enzyme-linked immunosorbent assay for human tau detection.

Immunoaffinity chromatography with a monoclonal antibody produced against bovine tau protein was used to purify tau proteins from human brain. Fifty grams of brain tissue yielded approximately 2 mg of pure tau proteins. The affinity-purified human tau was used to produce a high-titered rabbit anti-human tau serum. The monoclonal anti-tau antibody and the polyclonal rabbit anti-tau serum were then used to construct a sandwich enzyme-linked immunosorbent assay for detection of human tau proteins, with a sensitivity of 1 ng/ml.

Specific monoclonal antibodies against normal microtubule-associated protein-2 (MAP2) epitopes present in Alzheimer pathological structures do not recognize paired helical filaments.

We have developed monoclonal antibodies that detect normal microtubule-associated protein-2 (MAP2) epitopes in routinely fixed, paraffin-embedded tissue. The somatodendritic distribution of MAP2 in bovine and human nervous tissue was confirmed with several of these antibodies. Furthermore, some of these antibodies immunohistochemically labeled certain pathological structures in Alzheimer brain, especially neurites in senile plaques. Electron microscopic observations, however, indicate that these MAP2 epitopes are not located in the Alzheimer paired helical filaments themselves, but in amorphous granular structures coexistent with them. While the pathological nature of these structures is undetermined, they may represent artefactual modifications of normal cytoskeletal components.

Monoclonal antibodies with selective specificity for Alzheimer Tau are directed against phosphatase-sensitive epitopes.

A modified form of the microtubule-associated protein Tau is the major component of the paired helical filaments (PHF) found in Alzheimer's disease. The characterization of these posttranslational Tau modifications is hindered by the lack of sufficient PHF-Tau-specific markers. Here we describe several monoclonal antibodies, prepared by immunization with PHF, two of which showed a selective specificity for PHF-Tau without cross-reactivity with normal Tau. Epitope recognition by these two monoclonals was sensitive to alkaline phosphatase treatment. In Western blotting these monoclonal antibodies reacted specifically with the abnormally phosphorylated epitopes on Alzheimer's disease-associated PHF-Tau. One of the new antibodies can be used for the construction of a sandwich enzyme-linked immunosorbent assay for the specific detection of PHF-Tau without cross-reactivity to normal Tau proteins.

Radiolocalisation and imaging of stably HPLAP-transfected MO4 tumours with monoclonal antibodies and fragments.

Immunotargeting of PLAP-expressing tumours was studied for two radioiodinated, highly specific anti-PLAP monoclonal antibodies, 7E8 and 17E3, differing 10-fold in affinity, as well as for 7E8 F(ab')2 fragments. An anti-CEA monoclonal antibody or anti-CD3 F(ab')2 fragments were used as controls. Specific and non-specific targeting was examined in nude mice simultaneously grafted with PLAP-positive tumours derived from MO4 1-4 cells, and CEA-positive tumours, derived from 5583-S cells. Results indicated that (1) MO4 1-4 tumours, with a stable expression of PLAP on the plasma membrane, represent a useful new in vivo model for immunodirected tumour targeting; (2) differences in antibody affinity for PLAP in vitro are not reflected in antibody avidity for tumour cells in vivo; and (3) excellent selective and specific localisation of the PLAP-positive tumours is achieved when 7E8 F(ab')2 fragments are used. The high tumour/blood ratios (10.7 +/- 3.9 at 46 h after injection) were due to a much faster blood clearance of 7E8 F(ab')2 fragments. At this time point, the mean tumour/non-tumour tissue ratio was as high as 34.5, and the mean specific localisation index was 29.0. As expected, the F(ab')2 fragments provided high tumour imaging efficiency on gamma camera recording. These data imply important potentials of the PLAP/anti-PLAP system for immunolocalisation and therapy in patients, but also emphasise that in vitro criteria alone are not reflected in in vivo tumour localisation capacities of antibodies.