Hamiltonian Engineering with Multicolor Drives for Fast Entangling Gates and Quantum Crosstalk Cancellation.

Quantum computers built with superconducting artificial atoms already stretch the limits of their classical counterparts. While the lowest energy states of these artificial atoms serve as the qubit basis, the higher levels are responsible for both a host of attractive gate schemes as well as generating undesired interactions. In particular, when coupling these atoms to generate entanglement, the higher levels cause shifts in the computational levels that lead to unwanted ZZ quantum crosstalk. Here, we present a novel technique to manipulate the energy levels and mitigate this crosstalk with simultaneous off-resonant drives on coupled qubits. This breaks a fundamental deadlock between qubit-qubit coupling and crosstalk. In a fixed-frequency transmon architecture with strong coupling and crosstalk cancellation, additional cross-resonance drives enable a 90 ns CNOT with a gate error of (0.19±0.02)%, while a second set of off-resonant drives enables a novel CZ gate. Furthermore, we show a definitive improvement in circuit performance with crosstalk cancellation over seven qubits, demonstrating the scalability of the technique. This Letter paves the way for superconducting hardware with faster gates and greatly improved multiqubit circuit fidelities.

Molecular cloning of plane pollen allergen Pla a 3 and its utility as diagnostic marker for peach associated plane pollen allergy.

BACKGROUND: Non-specific lipid transfer proteins (nsLTP) are considered to provoke allergic symptoms to plane tree pollen, which are frequently associated with peach allergy. OBJECTIVE: The objective was to clone the cDNA of plane pollen nsLTP Pla a 3, to characterize IgE-binding and allergenic potency of recombinant Pla a 3 in comparison to its natural counterpart and peach nsLTP Pru p 3. METHODS: Natural Pla a 3 was purified from plane pollen and analysed by mass spectrometry (MS). Recombinant Pla a 3 was characterized by SDS-PAGE and CD spectroscopy. Specific IgE to extract, components of plane pollen and Pru p 3 was measured by ImmunoCAP in sera of patients allergic to either plane pollen (n = 10), peach (n = 15) or both (n = 15). Biological potency of the proteins was investigated by in vitro mediator release assays and IgE cross-reactivity by competitive ELISA. RESULTS: Two Pla a 3 isoforms were identified. Recombinant Pla a 3 showed high purity, structural integrity, IgE-binding capacity comparable to nPla a 3 and biological potency. Sensitization to plane pollen extract was confirmed in 24/25 plane pollen allergics. The frequency of sensitization to Pla a 3 was 53% among patients allergic to both plane pollen and peach and 10% among plane pollen allergics tolerating peach where most patients were sensitized to Pla a 1. Pla a 3 and Pru p 3 showed strong bi-directional IgE cross-reactivity in patients allergic to peach and plane pollen, but not in peach allergics tolerating plane pollen. Levels of IgE-binding were generally higher to Pru p 3 than to Pla a 3. CONCLUSION: Sensitization to Pla a 3 is relevant in a subgroup of plane pollen allergics with concomitant peach allergy. IgE testing with Pla a 3 may serve as a marker to identify plane pollen allergic patients at risk of LTP-mediated food reactions and thereby improve in vitro diagnostic procedures.

Pru p 3, the nonspecific lipid transfer protein from peach, dominates the immune response to its homolog in hazelnut.

BACKGROUND: Nonspecific lipid transfer proteins (nsLTPs) are important food allergens. Often, patients allergic to the nsLTP in peach suffer from allergy to hazelnuts. We aimed to analyse the T-cell response to Cor a 8, the nsLTP in hazelnut and its immunological cross-reactivity with the nsLTP in peach, Pru p 3. METHODS: Cor a 8-reactive T-cell lines (TCL) established from patients allergic to hazelnut and peach were stimulated with 12-mer peptides representing the complete amino acid sequence of Cor a 8 to identify its T-cell-activating regions and with Pru p 3 to investigate cellular cross-reactivity. T-cell clones specific for different major T-cell-activating regions of Pru p 3 were stimulated with Cor a 8. Both nsLTPs were subjected to endolysosomal degradation assays. Immunoglobulin E (IgE) cross-reactivity between Cor a 8 and Pru p 3 was assessed in inhibition enzyme-linked immunosorbent assay. RESULTS: No major T-cell-activating region was found among 26 T-cell-reactive peptides identified in Cor a 8. Although generated with Cor a 8, 62% of the TCL responded more strongly to Pru p 3. This cross-reactivity was mediated by T cells specific for the immunodominant region Pru p 3(61-75) . Peptide clusters encompassing this region were generated during lysosomal degradation of both nsLTPs. Cor a 8 was more rapidly degraded by lysosomal proteases than Pru p 3. Pre-incubation of sera with Pru p 3 completely abolished IgE binding to Cor a 8, which was not the case vice versa. CONCLUSIONS: T-cell reactivity to Cor a 8 is predominantly based on cross-reactivity with Pru p 3, indicating that the latter initiates sensitisation to its homolog in hazelnut. The limited allergenic potential of Cor a 8 seems to be associated with rapid lysosomal degradation during allergen processing and the lack of major T-cell-activating regions.

The non-specific lipid transfer protein, Ara h 9, is an important allergen in peanut.

BACKGROUND: Plant food allergy in the Mediterranean area is mainly caused by non-specific lipid transfer proteins (nsLTP). The aim of this study was to characterize peanut nsLTP in comparison with peach nsLTP, Pru p 3, and assess its importance in peanut allergy. METHODS: Peanut-allergic patients from Spain (n=32) were included on the basis of a positive case history and either a positive skin prick test or specific IgE to peanut. For comparison, sera of 41 peanut-allergic subjects from outside the Mediterranean area were used. Natural Ara h 9 and two isoforms of recombinant Ara h 9, expressed in Pichia pastoris, were purified using a two-step chromatographic procedure. Allergen characterization was carried out by N-terminal sequencing, circular dichroism (CD) spectroscopy, immunoblotting, IgE inhibition tests and basophil histamine release assays. RESULTS: Compared with natural peanut nsLTP, the recombinant proteins could be purified in high amounts from yeast supernatant (> or =45 mg/L). The identity of the proteins was verified by N-terminal amino acid sequencing and with rabbit nsLTP-specific antibodies. CD spectroscopy revealed similar secondary structures for all preparations and Pru p 3. The Ara h 9 isoforms showed 62-68% amino acid sequence identity with Pru p 3. IgE antibody reactivity to rAra h 9 was present in 29/32 Spanish and 6/41 non-Mediterranean subjects. Recombinant Ara h 9 showed strong cross-reactivity to nPru p 3 and similar IgE-binding capacity as nAra h 9. The two Ara h 9 isoforms displayed similar IgE reactivity. In peanut-allergic patients with concomitant peach allergy, Ara h 9 showed a weaker allergenic potency than Pru p 3 in histamine release assays. CONCLUSIONS: Ara h 9 is a major allergen in peanut-allergic patients from the Mediterranean area. Ara h 9 is capable of inducing histamine release from basophils, but to a lesser extent than Pru p 3.

Enhancement of hazelnut extract for IgE testing by recombinant allergen spiking.

BACKGROUND: Hazelnuts are a common cause of food allergic reactions. Most hazelnut allergic individuals in central and northern Europe are sensitized to Cor a 1, a member of the PR-10 protein family, while the lipid transfer protein Cor a 8 acts as a major allergen in the south of Europe. Other allergens, including profilin and seed storage proteins, may be important in subgroups of patients. Reliable detection of specific IgE in the clinical diagnosis of food allergy requires allergen reagents with a sufficient representation of all relevant allergen components. Some reported observations suggest that natural hazelnut extract may not be fully adequate in this respect. METHODS: The capacity of immobilized natural hazelnut extract to bind Cor a 1-, Cor a 2- and Cor a 8-specific IgE and IgG antibodies was investigated by serum adsorption and extract dilution experiments and by the use of allergen specific rabbit antisera. All measurements were performed with the ImmunoCAP assay platform. RESULTS: The experimental results revealed an incomplete capacity of immobilized hazelnut extract to capture IgE antibodies directed to the major allergen Cor a 1. Spiking of hazelnut extract with recombinant Cor a 1.04 prior to solid phase coupling gave rise to significantly enhanced IgE antibody binding from Cor a 1 reactive sera. The spiking did not negatively affect the measurement of IgE to extract components other than Cor a 1. CONCLUSION: A hazelnut allergen reagent with enhanced IgE detection capacity can be generated by supplementing the natural food extract with recombinant Cor a 1.04.

Identification of a plane pollen lipid transfer protein (Pla a 3) and its immunological relation to the peach lipid-transfer protein, Pru p 3.

BACKGROUND: An association between plane tree pollen allergy and plant food allergy has been described, but the cross-reacting allergens have not yet been identified. The aim of this study was the identification of homologous non-specific lipid-transfer proteins (nsLTPs) in plane pollen, and to investigate its immunological relationship with the peach LTP, Pru p 3. METHODS: Three different patient groups were recruited in Spain: 22 plane pollen-allergic patients without food allergy (A), 36 plane pollen-allergic patients with peach allergy (B) and 10 peach-allergic patients without plane pollen allergy (C). Proteins from plane pollen extract were fractionated by ion-exchange and reversed-phase chromatography. Further methods applied were N-terminal amino acid sequence analysis, immunoblotting, enzyme allergosorbent test, CAP and basophil histamine release assays. RESULTS: A 10 kDa IgE-reactive protein was purified from plane pollen and identified as nsLTP. Pla a 3 was characterized as a minor allergen (27.3%) in plane pollen-allergic patients without food allergy (A) and as a major allergen in plane pollen-allergic patients with peach allergy (B) showing a prevalence of IgE-reactivity of 63.8%. Group B contained patients sensitized to Pru p 3 without IgE-reactivity to plane-LTP (16.6%). By contrast, Pla a 3 IgE-reactive patients without sensitization to Pru p 3 could be found (16.6%). The sera of patients sensitized to both LTPs (50%), Pla a 3 and Pru p 3, showed different biological activity in histamine release assay: depending on individual patient's sera tested, Pla a 3 showed a similar, a stronger or a weaker allergenic potency in comparison with Pru p 3. CONCLUSIONS: Plane LTP is a major allergen in plane pollen-allergic patients with peach allergy recruited in the Mediterranean area. The results of histamine release tests and different IgE-binding profiles pointed towards the existence of species-specific IgE epitopes. Likewise, no general conclusion on the sensitizer could be made.

The value of FDG-PET and bone scintigraphy with SPECT in the primary diagnosis and follow-up of patients with chronic osteomyelitis of the mandible.

OBJECTIVE: To appraise the value of FDG-PET and bone scintigraphy using SPECT in the primary diagnosis and follow-up of patients with chronic osteomyelitis of the mandible (COM). METHODS: In a prospective study the pattern of tracer uptake was investigated using 2 diagnostic methods in 42 patients. Results were compared with histology and radiographs as well as clinical and laboratory parameters. RESULTS: The use of FDG-PET in the primary diagnosis of COM resulted in a sensitivity of 64% and a specificity of 77.7%. The sensitivity of SPECT was 84% and the specificity 33.3%. During the follow-up period of these patients the sensitivity of SPECT increased to 93.7%, while the specificity decreased (6.6%). The sensitivity and specificity of FDG-PET for this follow-up group were 62.5 and 80%, respectively. CONCLUSION: Because of its high sensitivity, SPECT is vastly superior to other diagnostic methods in initiating treatment. In the follow-up period it might be replaced by FDG-PET, which reflects the disease course better and indicates the time of clinical remission.

Structural, immunological and functional properties of natural recombinant Pen a 1, the major allergen of Brown Shrimp, Penaeus aztecus.

BACKGROUND: Recombinant allergens are considered the basis for new diagnostic approaches and development of novel strategies of allergen-specific immunotherapy. As Pen a 1 from brown shrimp Penaeus aztecus is the only major allergen of shrimp and binds up to 75% of all shrimp-specific IgE antibodies this molecule may be an excellent model for the usage of allergens with reduced IgE antibody-binding capacity for specific immunotherapy. AIM: The aim was to clone, express and characterize a full-length recombinant Pen a 1 molecule and compare it with natural Pen a 1 in regard to structural and immunological parameters such as IgE antibody capacity and ability to induce IgE-mediated mediator release. METHODS: Total RNA was isolated from P. aztecus and a rapid amplification of cDNA ends (5' RACE) was performed to obtain full-length cDNA coding for Pen a 1. Using a gene-specific primer, PCR was performed and full-length cDNA was cloned and sequenced. Recombinant His-tagged Pen a 1 was isolated from Escherichia coli under native conditions by immobilized metal affinity chromatography. Secondary structure of natural and recombinant Pen a 1 was compared by circular dichroism (CD) spectroscopy, and the IgE antibody-binding capacity evaluated by RAST. The allergenic potency was tested by the capability of natural and recombinant Pen a 1 to induce mediator release in a murine and human in vitro model of IgE-mediated type I allergy. RESULTS: The deduced amino-acid sequence was 284 residues long and amino-acid sequence identities with allergenic and non-allergenic tropomyosins ranged from 80% to 99% and 51% to 58%, respectively. The analysis of the secondary structure of natural and recombinant Pen a 1 by CD spectroscopic analysis showed that both nPen a 1 and rPen a 1 had alpha-helical conformation that is typical for tropomyosin. The IgE antibody binding capacities of nPen a 1 and r Pen a1 were found to be essentially identical by RAST. The mediator release experiments using both wild-type and humanized rat basophilic leukaemia 30/25 cells showed that rPen a 1 and nPen a 1 induced a similar level of mast cell activation. CONCLUSIONS: Recombinant Pen a 1 and natural Pen a 1 are structurally and immunologically identical and rPen a 1 may be used as the basis for component-resolved diagnosis and the generation of modified shrimp tropomyosin for allergen-specific immunotherapy. The results of the animal studies indicate that C3H/HeJ mice that were sensitized with shrimp extract in combination with cholera toxin as adjuvant may be a suitable model to study shrimp allergy.

Re-distribution of tenascin-C in the parotid acinar cells. An early marker of radiation-induced damage of salivary glands?

PURPOSE: In order to investigate the early changes in the expression of tenascin-C, following irradiation and the associated functional impairment of salivary glands. MATERIALS AND METHODS: Fifteen rabbits were used for the study. Five provided control parotid gland tissue and a further 10 rabbits were scintigraphically examined prior to and 24 h after 15/30 Gy. Glands were studied histologically using HE-staining and tenascin-C antibodies. RESULTS: Reduction in the salivary ejection fraction (SEF) was observed in all irradiated glands. Simultaneously, a marked re-distribution of tenascin-C expression was noticed. Reactivity detected in the intercalated, secretory ducts and perineurinal regions prior to radiation was noticed intracellularly after 24 h. Furthermore, nerves showed tenascin-C expression in the Schwann cells, but no longer perineurinally. Myofibroblasts were also observed in the stroma. CONCLUSION: This study proves the ability to predict functional disorders of salivary function as early as 24 h after radiation and provides evidence of the participation of tenascin-C in the pathological process of radiation-induced damage in salivary glands.

Early immunohistochemical and functional markers indicating radiation damage of the parotid gland.

In order to evaluate the correlation between functional impairment and changes in the expression pattern of immunohistochemical antibodies in the early phase of radiation-induced dysfunction of salivary glands, eight rabbits were scintigraphically examined prior to and 24 h after irradiation with 15 Gy. The parotid glands were studied using HE-staining, Ki-67, alpha-smooth muscle actin (ASMA) and tenascin-C antibodies at every scintigraphic examination. The results demonstrated a significant alteration in the 99mTc-pertechnetate uptake in all irradiated glands. HE-staining showed no relevant impairment of salivary gland tissue in this early phase. Immunohistochemically, we observed a marked re-distribution of ASMA and tenascin-C as well as a reduction of the proliferating rate of acinar cells. This immunohistochemical change correlated with the functional impairment manifested scintigraphically. This study proves the possibility to assess disorders of salivary gland function with immunohistological antibodies as early as 24 h after irradiation and yields the prerequisites to prove the effects of radioprotective agents on salivary gland tissues.

Scintigraphic evaluation of lacrimal glands using a rabbit experimental model.

The aim of this study was to establish an experimental model to evaluate functional changes in lacrimal gland parenchyma using gamma scintigraphy. Although the lacrimal glands of the rabbit have frequently been used for ophthalmological research, scintigraphic evaluation of these glands has received far less attention and we could not find any reports concerning this topic in the literature. Ten rabbits were used for the study; in 4 of them, the orbital region was dissected to provide the topographic anatomy of the lacrimal glands. Four rabbits underwent a static scintigraphy after excision of a unilateral lacrimal gland. Changes in the pattern of tracer uptake indicated the exact position of the gland on the scintiscan. One rabbit served as a control, and another one was used to prove the surgical accessibility of the gland. Using a frontal projection of the head the (99m)TcO(-)(4) uptake of the rabbit lacrimal glands could be identified and evaluated in the upper lateral region of the scintiscan. In conclusion, the lacrimal glands of the rabbit provide an appropriate experimental model to study quantitative disturbances of lacrimal secretion using scintigraphy and enable the assignment of functional impairment to morphological changes of the lacrimal parenchyma.

The superficial mandibular gland of the rabbit: a new experimental model for scintigraphic evaluation of salivary glands.

The aim of this study was to provide an appropriate experimental model to study functional changes in salivary glands using scintigraphy. Although the rabbit was frequently used for laboratory experiments, there are only a few studies that describe the exact position of its different salivary glands on the sialoscintigram. Twenty rabbits were used for the study; ten of them were anatomically dissected to provide the required topographic anatomy on the different salivary and lacrimal glands. The remaining ten animals underwent a static scintigraphy after extirpation of a particular salivary gland. Changes in the pattern of tracer uptake indicated the exact position of every gland allowing its evaluation. The results show that the 99mTcO4- uptake can only be selectively evaluated in two salivary glands, the superficial mandibular gland and the parotid gland. The superficial mandibular gland was proved to be a particularly useful model to evaluate functional changes of salivary gland parenchyma due to its well defined and high enhanced structure which allows a precise detection and measurement of the tracer uptake. Additionally, the good surgical accessibility of this gland and the existence of a well defined capsule facilitate associated histological studies of its parenchyma.

Single photon emission tomography: scintigraphy in the assessment of mandibular invasion by head and neck cancer.

BACKGROUND: This study was performed to determine the effectiveness of single photon emission tomography (SPET) in contributing to the initial staging of patients with cancers of the head and neck because information about osseous infiltration of head and neck cancer is of major importance for staging and planning of treatment. METHODS: A retrospective analysis of 89 cases with probable tumor infiltration of the mandible by oral/pharyngeal cancer was undertaken by comparing the preoperative SPET as well as CT-scans, if available, with clinical and postoperative histological examinations. RESULTS: For SPET examinations, a sensitivity of 95% and a specificity of 48% were calculated. The positive predictive value was 65%, and the negative predictive value was 93%. CT scans showed a sensitivity of 75% and a specificity of 78%. The positive predictive value was 65%, and the negative predictive value was 93%. CONCLUSIONS: SPET is a powerful method to detect infiltration of carcinomas into the head and neck skeleton. Its sensitivity is high and superior to CT scans, although the specificity is small due to a high number of false positive results.

Assessment of the viability of microvascularized bone grafts after mandibular reconstruction by means of bone SPET and semiquantitative analysis.

The purpose of this study was to investigate the value of bone scintigraphy including single-photon emission tomography (SPET) and semiquantitative analysis for the assessment of graft viability following microvascularized bone transplantation. We evaluated 60 scintigraphic studies of 36 patients with 39 bone grafts. Thirty-four investigations were performed 6-11 days (early bone scans) and 26 up to 11 months (late bone scans) after mandibular reconstruction. After administration of 550 MBq technetium-99m methylene diphosphonate, planar scintigrams and a SPET study were performed. The data were reconstructed iteratively. Scans were evaluated visually and semiquantitatively by a region of interest technique using the ratio between transplant and cranium (T/C). Patients with uncomplicated healing showed a T/C ratio >1.0 in early and late bone scans. In cases with necrosis, the T/C ratio was below 1.0 when performing early bone scans. However, in late bone scans, some patients with necrosis showed a slightly increased uptake and a T/C ratio >1.0. The data demonstrate that as early as 6-11 days after mandibular reconstruction, increased tracer uptake proves that the surgery has been successful and indicates a normal healing process. Especially in the early bone scans no false-positive or false-negative results were observed and the T/C ratio clearly differentiated between vital and non-vital bone grafts. At later times false-positive findings could be observed; these were, however, rare because of the significantly higher tracer uptake of the healthy grafts when compared with completely or partially necrotic transplants.

Microvascular submandibular gland transfer for severe cases of keratoconjunctivitis sicca.

Free submandibular salivary gland transfer was investigated as a surgical method for the treatment of severe keratoconjunctivitis sicca. In an animal model, we examined the tolerance of warm ischemia of the submandibular gland. After temporary interruption of the blood supply (1 to 6 hours), the morphologic changes in the submandibular gland were analyzed histologically and immunohistochemically in 41 rabbits. From 1.5 hours ischemia onward, an increasing structural damage of the parenchyma with emphasis on the secretory cells was seen. Six hours of ischemia caused total necrosis of the salivary gland. Our clinical experience includes 24 highly selected patients suffering from keratoconjunctivitis sicca, in whom we transferred 31 autologous submandibular glands to the temple for permanent autologous tear substitution within the past 4 years. The glands were implanted into a pocket prepared in the temporalis muscle, and the nourishing vessels were anastomosed to the superficial temporal artery and vein. The submandibular duct was implanted into the upper lateral conjunctival fornix. The transferred glands were left denervated. In addition to the clinical examination, scintigraphy with Tc 99m pertechnetate was used to document the graft's viability after the transfer. Viable incorporation with longstanding secretory function occurred in 26 of the 30 transplanted denervated salivary glands. The resulting lubrication of the treated eyes was irregular for up to 3 months in almost even case. One year after surgery, all patients with a viable transplant developed at least occasional epiphora, which was surgically managed by reducing the size of the graft in 10 patients. No severe side effects were seen in this series. The ophthalmologic evaluation of the method included the assessment of dry eye symptoms and of the volume and quality of ocular lubrication (Schirmer test, fluorescein break-up time), the pathology of the ocular surface (rose bengal staining), and the need for pharmaceutical tear substitutes. One year after surgery, 18 of 27 cases assessed were judged as significantly improved by these tests.

Radioiodine therapy in Graves' hyperthyroidism: determination of individual optimum target dose.

UNLABELLED: The aim of this study was to investigate the results of different volume-dependent target doses on clinical outcome 6 months after radioiodine therapy (RITh) and its correlation with post therapeutic thyroid volumes (Vpost) in patients with Graves' disease. This analysis was designed to determine factors improving the results of radioiodine therapy without increasing target doses generally, as has been recommended recently. We studied consecutive data from 102 patients with Graves' disease, who had initial radioiodine therapy between 1991 and 1995. The 131I activities were calculated according to the formula of Marinelli. In addition to the normal calculation individual target doses were adjusted to the thyroid volumes of each patient before therapy. For statistical evaluation, the patients were divided into three subgroups of comparable sample sizes: Group I included those with a thyroid volume <15 ml before therapy. Group II included those ranging from a 15-25 ml volume before therapy and group III included those with thyroid volumes >25 ml. Laboratory thyroid parameters and thyroid volumes were measured in those groups before and 6 months after therapy. RESULTS: Analysis of all patients revealed a significantly higher rate of hypothyroidism (54%) and fewer cases of hyperthyroidism (15%) six months after therapy in cases with Vpost smaller than 8 ml. The median Vpost needed to achieve an optimum therapeutic success rate (rate of eu- or hypothyroidism) was smaller than 5 ml in group I and smaller than 10 ml in group II. Therapeutic success was associated with different target doses in each group, 150,220 and 260 Gy for groups I, II, and III respectively. CONCLUSIONS: Post therapeutic thyroid volumes correlated significantly with clinical outcome six months after therapy. An adjustment of the target doses based on thyroid volumes before therapy will lead to an appropriate reduction of thyroid volumes. Thus, in the individual case clinical outcome could be improved without applying higher target doses in all patients. This would ensure a better utilization of limited resources in medical care e.g. through a shorter hospital stay.

Quality of salivary tears following autologous submandibular gland transplantation for severe dry eye.

BACKGROUND: This study aimed to characterise the composition of the pre-ocular fluid after transplantation of the autologous submandibular gland (SMG) for patients with severe dry eye. METHODS: Stimulated and unstimulated pre-ocular fluid from 15 patients (17 eyes) with a viable SMG graft ("SMG-salivary tears"), as well as normal tears and SMG saliva (20 normal subjects/ 20 eyes), was sampled. As global tear parameters, fern pattern analysis and SDS gel electrophoresis were performed. As specific quality parameters, total protein content, secretory immunoglobulin A (SIgA), lysozyme, amylase, sodium, potassium and osmolality were measured using routine laboratory methods. The flow rate of SMG-salivary tears was determined in 5 patients by means of sequential scintillography. RESULTS: The fern pattern of SMG-salivary tears was coarse and thus more similar to normal SMG saliva than tears. SDS gel electrophoresis of the SMG-salivary tears showed albumin and two unidentified proteins in addition to the normal tear pattern. Osmolality and total protein content of SMG-salivary tears were higher than in normal SMG saliva, but still lower than in normal tears. High activities of normal tear antibacterial proteins (SIgA, lysozyme and amylase) were detected in the salivary tears. Stimulation of the secretion did not alter the composition of SMG-salivary tears. The flow rate of SMG-salivary tears was closer to that of normal tears than normal SMG saliva. CONCLUSION: Salivary tears resulting from SMG-transplantation represent condensed SMG saliva. Thus their quality is intermediate between normal tears and normal SMG saliva. High levels of secretory proteins demonstrate that the gland maintains an active function. Surgical denervation and residual tear components from the ocular surface are the most likely factors to cause the complex differences between normal SMG saliva and SMG-salivary tears. The effects of this secretion on the ocular surface are currently being evaluated in a clinical and laboratory study.

New approach for separating Bacillus subtilis metalloprotease and alpha-amylase by affinity chromatography and for purifying neutral protease by hydrophobic chromatography.

Proteases are commonly used in the biscuit and cracker industry as processing aids. They cause moderate hydrolysis of gluten proteins and improve dough rheology to better control product texture and crunchiness. Commercial bacterial proteases are derived from Bacillus fermentation broth. As filtration and ultrafiltration are carried out as the only recovery steps, these preparations contain also alpha-amylase and beta-glucanase as the main side activities. The aim of this study is to purify and characterize the Bacillus subtilis metalloprotease from a commercial preparation, in order to study separately the impact of the protease activity with regards to its functionality on biscuit properties. Purification was achieved by means of affinity chromatography on Cibacron Blue and HIC as a polishing step. Affinity appeared to be the most appropriate matrix for large scale purification while ion exchange chromatography was inefficient in terms of recovery yields. The crude product was first loaded on a Hi Trap Blue column (34 microm, Pharmacia Biotech); elution was carried out with a gradient of NaCl in the presence of 1 mM ZnCl2. This step was only efficient in the presence of Zn cations, because this salt promoted both protease stabilization resulting in high recovery yields and also complexation of amylase units into dimers resulting in amylase retention on the column and a better separation of the 3 activities. Beta-glucanase was mostly non retained on the column and a part was coeluted with the protease. This protease fraction was then loaded on a Resource Phe column (15 microm, Pharmacia Biotech) in a last step of polishing. Elution was carried out with a linear gradient of 100-0% ammonium sulfate 1.3 M; protease was eluted at the beginning of the gradient and well separated from amylase and glucanase trace impurities. The homogeneity of the purified protease was confirmed by SDS-PAGE, which showed that its MW was about 38. pH and temperature optima were also determined on the fraction.

Quality of salivary tears following autologous submandibular gland transplantation for severe dry eye.

BACKGROUND: This study aimed to characterise the composition of the pre-ocular fluid after transplantation of the autologous submandibular gland (SMG) for patients with severe dry eye. METHODS: Stimulated and unstimulated pre-ocular fluid from 15 patients (17 eyes) with a viable SMG graft ("SMG-salivary tears"), as well as normal tears and SMG saliva (20 normal subjects/20 eyes), was sampled. As global tear parameters, fern pattern analysis and SDS gel electrophoresis were performed. As specific quality parameters, total protein content, secretory immunoglobulin A (SIgA), lysozyme, amylase, sodium, potassium and osmolality were measured using routine laboratory methods. The flow rate of SMG-salivary tears was determined in 5 patients by means of sequential scintillography. RESULTS: The fern pattern of SMG-salivary tears was coarse and thus more similar to normal SMG saliva than tears. SDS gel electrophoresis of the SMG-salivary tears showed albumin and two unidentified proteins in addition to the normal tear pattern. Osmolality and total protein content of SMG-salivary tears were higher than in normal SMG saliva, but still lower than in normal tears. High activities of normal tear antibacterial proteins (SIgA, lysozyme and amylase) were detected in the salivary tears. Stimulation of the secretion did not alter the composition of SMG-salivary tears. The flow rate of SMG-salivary tears was closer to that of normal tears than normal SMG saliva. CONCLUSION: Salivary tears resulting from SMG-transplantation represent condensed SMG saliva. Thus their quality is intermediate between normal tears and normal SMG saliva. High levels of secretory proteins demonstrate that the gland maintains an active function. Surgical denervation and residual tear components from the ocular surface are the most likely factors to cause the complex differences between normal SMG saliva and SMG-salivary tears. The effects of this secretion on the ocular surface are currently being evaluated in a clinical and laboratory study.

Influence of radioiodine therapy on urinary iodine excretion.

AIM: In 214 patients with benign thyroid diseases the time-course of urinary iodine excretion (UIE) was investigated in order to identify changes after radioiodine therapy (RITh). METHOD: UIE was measured photometrically (cerium-arsenite method) and related to urinary creatinine on the first and last day of the radioiodine test and then three days, seven days, four weeks, and six months after 131I administration. RESULTS: As compared with the level found immediately before radioiodine therapy, median UIE had almost doubled four weeks after therapy and was still significantly elevated six months after therapy. This increase correlated significantly with the target volume as measured by scintigraphy and sonography. CONCLUSIONS: The persistent elevation of UIE for months after RITh is a measure of treatment-induced damage to thyrocytes. Therefore, in view of the unfavourable kinetics of iodine that follow it, RITh should if possible be given via a single-dose regime.