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Size segregation in granular materials is a universal phenomenon popularly known as the Brazil nut effect (BNE), from the tendency of larger nuts to end on the top of a shaken container. In nature, fast granular flows bear many similarities with well-studied mixing processes. Instead, much slower phenomena, such as the accumulation of ferromanganese nodules (FN) on the seafloor, have been attributed to the BNE but remain essentially unexplained. Here we document, for the first time, the BNE on sub-millimetre particles in pelagic sediment and propose a size segregation model for the surface mixed layer of bioturbated sediments. Our model explains the size distribution of FN seeds, pointing to a uniform segregation mechanism over sizes ranging from < 1 mm to > 1 cm, which does not depend on selective ingestion by feeding organisms. In addition to explaining FN nucleation, our model has important implications for microfossil dating and the mechanism underlying sedimentary records of the Earth's magnetic field., (© 2022. The Author(s).)

Sampling of sediment cores using plastic U-channels has made possible the acquisition of detailed records of paleomagnetic secular variation, geomagnetic polarity, environmental magnetic studies, and relative paleointensity over the past several million years. U-channel measurements provide the great advantage of rapid measurements of long sediment cores, but the signal resolution is attenuated by the response function of the magnetometer sensors, which therefore restrains the recovery of rapid and large-amplitude field changes. Here we focus on the suitability of the dynamics of reversals and excursions derived from U-channel measurements. We compare successive individual paleomagnetic directions of 1.5 cm × 1.5 cm × 1.5 cm cubic discrete samples with those of a 1.5-m equivalent U-channel sample train obtained by placing the samples adjacent to each other. We use varying excursion and transition lengths and generate transitional directions that resemble those of the most detailed paleomagnetic records. Excursions with opposite polarity directions recorded over less than 7.5 cm are barely detected in U-channel measurements. Regarding reversals, U-channel measurements smooth the signal of low-resolution records and generate artificial transitional directions. Despite producing misleading similarities with the overall structure of transition records, longer transitional intervals fail also to reproduce the complexity of field changes. Finally, we test the convolution of magnetization by different response functions. The simulation reveals that even small response function changes can generate significant differences in results.

Paleomagnetic directional data were obtained from fourteen 0 to 2 Ma old lava flows at Basse-Terre Island (Guadeloupe, French West Indies). Five reversed polarity flows are consistent with their Matuyama age between 1.6-1.5 Ma and 875-790 ka while the ages of the other nine normal polarity units tie them to the Olduvai subchron and the Brunhes Chron. These directions have been combined with previous results obtained from Basse-Terre Island. The overall mean direction (D = -1.2°, I = 31.4°, α 95  = 3.3°) obtained from the 39 non-transitional flows from Basse-Terre Island is indistinguishable from the expected geocentric axial dipole value (D = 0°, I = 29.8°). The dispersion measured from the angular standard deviation of the Virtual Geomagnetic Poles (VGPs) was found to be close to, but smaller than the predictions of geomagnetic models. Together with further directions from the nearby Martinique Island, the 45 directions obtained within the Brunhes chron provide the most robust estimate of the statistical distribution of paleosecular variation (PSV) at this latitude. The sequence of directions shows episodes of high amplitude secular variation that are coeval with several geomagnetic events including the last reversal documented by five transitional directions. Finally, three lava flows have recorded a transitional behavior which could be link to two excursions, the Laguna del Sello (at ~340 ka) and the Pringle Falls (at ~210 ka) events.

In this study, biologically synthesized iron oxide nanoparticles, called magnetosomes, are made fully biocompatible by removing potentially toxic organic bacterial residues such as endotoxins at magnetosome mineral core surfaces and by coating such surface with poly-L-lysine, leading to magnetosomes-poly-L-lysine (M-PLL). M-PLL antitumor efficacy is compared with that of chemically synthesized iron oxide nanoparticles (IONPs) currently used for magnetic hyperthermia. M-PLL and IONPs are tested for the treatment of glioblastoma, a dreadful cancer, in which intratumor nanoparticle administration is clinically relevant, using a mouse allograft model of murine glioma (GL-261 cell line). A magnetic hyperthermia treatment protocol is proposed, in which 25 µg in iron of nanoparticles per mm 3 of tumor are administered and exposed to 11 to 15 magnetic sessions during which an alternating magnetic field of 198 kHz and 11 to 31 mT is applied for 30 minutes to attempt reaching temperatures of 43-46 °C. M-PLL are characterized by a larger specific absorption rate (SAR of 40 W/g Fe compared to 26 W/g Fe for IONPs as measured during the first magnetic session), a lower strength of the applied magnetic field required for reaching a target temperature of 43-46 °C (11 to 27 mT compared with 22 to 31 mT for IONPs), a lower number of mice re-administered (4 compared to 6 for IONPs), a longer residence time within tumours (5 days compared to 1 day for IONPs), and a less scattered distribution in the tumour. M-PLL lead to higher antitumor efficacy with full tumor disappearances achieved in 50% of mice compared to 20% for IONPs. This is ascribed to better ability of M-PLL, at equal iron concentrations, to maintain tumor temperatures at 43-46°C over a longer period of times., Competing Interests: Competing Interests: The authors have declared that no competing interest exists.

Geomagnetic dipole moment variations associated with polarity reversals and excursions are expressed by large changes of the cosmogenic nuclide beryllium-10 ( 10 Be) production rates. Authigenic 10 Be/ 9 Be ratios (proxy of atmospheric 10 Be production) from oceanic cores therefore complete the classical information derived from relative paleointensity (RPI) records. This study presents new authigenic 10 Be/ 9 Be ratio results obtained from cores MD05-2920 and MD05-2930 collected in the west equatorial Pacific Ocean. Be ratios from cores MD05-2920, MD05-2930 and MD90-0961 have been stacked and averaged. Variations of the authigenic 10 Be/ 9 Be ratio are analyzed and compared with the geomagnetic dipole low series reported from global RPI stacks. The largest 10 Be overproduction episodes are related to dipole field collapses (below a threshold of 2 × 10 22  Am 2 ) associated with the Brunhes/Matuyama reversal, the Laschamp (41 ka) excursion, and the Iceland Basin event (190 ka). Other significant 10 Be production peaks are correlated to geomagnetic excursions reported in literature. The record was then calibrated by using absolute dipole moment values drawn from the Geomagia and Pint paleointensity value databases. The 10 Be-derived geomagnetic dipole moment record, independent from sedimentary paleomagnetic data, covers the Brunhes-Matuyama transition and the whole Brunhes Chron. It provides new and complementary data on the amplitude and timing of millennial-scale geomagnetic dipole moment variations and particularly on dipole moment collapses triggering polarity instabilities.

Polarity reversals of the geomagnetic field are a major feature of the Earth's dynamo. Questions remain regarding the dynamical processes that give rise to reversals and the properties of the geomagnetic field during a polarity transition. A large number of paleomagnetic reversal records have been acquired during the past 50 years in order to better constrain the structure and geometry of the transitional field. In addition, over the past two decades, numerical dynamo simulations have also provided insights into the reversal mechanism. Yet despite the large paleomagnetic database, controversial interpretations of records of the transitional field persist; they result from two characteristics inherent to all reversals, both of which are detrimental to an ambiguous analysis. On the one hand, the reversal process is rapid and requires adequate temporal resolution. On the other hand, weak field intensities during a reversal can affect the fidelity of magnetic recording in sedimentary records. This paper is aimed at reviewing critically the main reversal features derived from paleomagnetic records and at analyzing some of these features in light of numerical simulations. We discuss in detail the fidelity of the signal extracted from paleomagnetic records and pay special attention to their resolution with respect to the timing and mechanisms involved in the magnetization process. Records from marine sediments dominate the database. They give rise to transitional field models that often lead to overinterpret the data. Consequently, we attempt to separate robust results (and their subsequent interpretations) from those that do not stand on a strong observational footing. Finally, we discuss new avenues that should favor progress to better characterize and understand transitional field behavior.

No consensus has been reached so far on the properties of the geomagnetic field during reversals or on the main features that might reveal its dynamics. A main characteristic of the reversing field is a large decrease in the axial dipole and the dominant role of non-dipole components. Other features strongly depend on whether they are derived from sedimentary or volcanic records. Only thermal remanent magnetization of lava flows can capture faithful records of a rapidly varying non-dipole field, but, because of episodic volcanic activity, sequences of overlying flows yield incomplete records. Here we show that the ten most detailed volcanic records of reversals can be matched in a very satisfactory way, under the assumption of a common duration, revealing common dynamical characteristics. We infer that the reversal process has remained unchanged, with the same time constants and durations, at least since 180 million years ago. We propose that the reversing field is characterized by three successive phases: a precursory event, a 180° polarity switch and a rebound. The first and third phases reflect the emergence of the non-dipole field with large-amplitude secular variation. They are rarely both recorded at the same site owing to the rapidly changing field geometry and last for less than 2,500 years. The actual transit between the two polarities does not last longer than 1,000 years and might therefore result from mechanisms other than those governing normal secular variation. Such changes are too brief to be accurately recorded by most sediments.

We show that a model, recently used to describe all the dynamical regimes of the magnetic field generated by the dynamo effect in the von Kármán sodium experiment, also provides a simple explanation of the reversals of Earth's magnetic field, despite strong differences between both systems. The validity of the model relies on the smallness of the magnetic Prandtl number.

Independent records of relative magnetic palaeointensity from sediment cores in different areas of the world can be stacked together to extract the evolution of the geomagnetic dipole moment and thus provide information regarding the processes governing the geodynamo. So far, this procedure has been limited to the past 800,000 years (800 kyr; ref. 3), which does not include any geomagnetic reversals. Here we present a composite curve that shows the evolution of the dipole moment during the past two million years. This reconstruction is in good agreement with the absolute dipole moments derived from volcanic lavas, which were used for calibration. We show that, at least during this period, the time-averaged field was higher during periods without reversals but the amplitude of the short-term oscillations remained the same. As a consequence, few intervals of very low intensity, and thus fewer instabilities, are expected during periods with a strong average dipole moment, whereas more excursions and reversals are expected during periods of weak field intensity. We also observe that the axial dipole begins to decay 60-80 kyr before reversals, but rebuilds itself in the opposite direction in only a few thousand years.

A fragment of the human lipoprotein lipase (LPL) cDNA (405 bp, 5' terminal end) was cloned in an expression vector to produce a approximately 17 kDa fusion peptide and was used as antigen to produce a high titre anti-LPL monoclonal antibody (10C3 MAb). This antibody reacts with both native and denatured forms of LPL from different tissue and animal sources. Competition studies with heparin indicate that 10C3 MAb is specific for an epitope at a heparin binding site. The antibody does not inhibit LPL enzyme activity, indicating that the antigenic epitope is not situated within or in the proximity of the LPL catalytic region. With these characteristics, 10C3 MAb should prove to be a useful immunochemical tool in clinical as well as in fundamental investigations on the metabolism of triglyceride-rich lipoproteins and in studies on the functional anatomy of LPL.

IFAPa-400, a 400-kDa developmentally regulated protein thought to be associated with intermediate filaments, has been purified from chick embryo hearts to investigate its interaction with vimentin and other IF proteins and to identify other cellular components to which this cytoskeletal protein associates. Previous studies suggested that this protein was associated with the vimentin-containing intermediate filament lattice of myoblasts and neuroblasts before their terminal differentiation, providing these cells with a particular intermediate filament cytoskeleton that could satisfy specific mechanical requirements during their intense morphogenetic activities. Although IFAPa-400 partially reassociated with vimentin and desmin in disassembly-reassembly experiments using crude IF preparations from chick embryo hearts, in vitro recombination of purified IFAPa-400 with vimentin and desmin failed to demonstrate any direct association. When purified IFAPa-400 was used as a probe in blot overlay assays, however, specific binding to vimentin and desmin was observed, providing the first evidence of a physical association between IFAPa-400 and intermediate filament proteins. The blot overlay experiments also demonstrated that IFAPa-400 binds to two unidentified polypeptides of 19 and 32 kDa. These results are thus consistent with the hypothesis that a structural lattice requiring a vimentin-IFAPa-400 combination constitutes the intermediate filament system of myogenic and neurogenic cells.

A variety of techniques have been used to identify the amino acid residues of bacterial superantigens involved in their interactions with major histocompatibility complex (MHC) class II and T-cell receptor (TCR). In this study, we isolated a naturally mutated staphylococcal enterotoxin A (SEA) from three different Staphylococcus aureus strains, in which the amino acid at position 60 has been changed from aspartic acid (D) to asparagine (N). We then studied the influence of this change on the immunological activities of SEA. Our results demonstrated that this mutation does not affect the capacity of SEA to bind MHC class II molecules and consequently activates human monocytes and peripheral blood lymphocytes. In contrast, mutated SEA failed to stimulate the proliferation of murine splenic lymphocytes of two different strains, and when presented by human MHC class II molecules, it also failed to activate murine cell line 3DT, which expresses the SEA-specific TCR V beta element (V beta 1). These results indicate that this mutation alters the interaction between SEA and murine TCR. The reactivity patterns of the mutated SEA with two specific anti-SEA monoclonal antibodies suggested that the observed effect of the isolated mutation in the murine system might be due to certain conformational changes in the SEA molecule introduced upon changing the D at position 60 to N. Site-directed mutagenesis of the N residue to D or to glycine reconstituted the ability of SEA to stimulate murine splenic lymphocytes. The different effects of this natural mutation at position 60 on the immunological activities of SEA with murine and human cells highlight the relevance of the affinity and avidity in SEA-TCR interactions in the function of different species or may reflect a difference in epitope specificity.

To determine differences in the patterns of expression of Drosophila small heat shock proteins (shsp) during normal development in the absence of stress, proteins obtained from head, thorax and gonads of young (0-12 h, 3 days), middle-aged (3-6 days) and 15- to 20-day-old adult flies were separated on SDS-PAGE gels and blotted with monoclonal antibodies against hsp23 and hsp26. hsp23 was found in the heads and gonads of young males and females. In contrast, the maximum expression of hsp26 was seen in gonads of young flies, and it was only lightly detected in the brain. The expression of both proteins decreased as flies aged. This age-related decrease was particularly striking for hsp23 in females. The immunoblot results obtained were complemented by immunostaining of thin parasagittal sections of whole fly bodies. Hsp23 was found to be expressed in the brain, thoracic ganglion, fat body and gonads of young (0-12 h) males and females. On the other hand, hsp26 was essentially detected in ovaries and testes of these young flies. The analysis of the tissue expression of both proteins demonstrate that each shsp has a distinct cellular localization. In the central nervous system, hsp23 and hsp26 were present in the neurocytes of the brain and the thoracic ganglion. In addition, hsp23 (but not hsp26) was also detected in the central neuropile of these two organs. In testis, hsp26 was localized in the cytoplasm of spermatocytes and, probably, in the spermatid bundles. In contrast, hsp23 was detected at the periphery of cells (membranes).(ABSTRACT TRUNCATED AT 250 WORDS)

Segment polarity genes define the cell states that are required for proper organization of each metameric unit of the Drosophila embryo. Among these, the gooseberry locus has been shown to be composed of two closely related genes which are expressed in an overlapping single-segment periodicity. We have used specific antibodies raised against the protein product of the gooseberry proximal (gsb-p) gene to determine the spatial distribution of this antigen in wild type embryos, and to monitor the effects of segment polarity mutants on the pattern of the gsb-p protein distribution. We find that the gsb-p protein accumulates beneath each posterior axonal commissure in the progeny of neuroblasts deriving from the epidermal compartments of wingless (wg) and engrailed (en) expression. The results of this analysis support the idea that gsb-p has a specific role in the control of cell fates during neurogenesis, and indicate that en and wg provide critical positional cues to define the domain in which gsbp will be activated. Furthermore, these data suggest that, in order to be expressed in the embryonic CNS, gsb-p may preliminarily require activity of the gooseberry-distal gene in the epidermis.

Hereditary tyrosinemia is characterized by a deficiency of the enzyme fumarylacetoacetate hydrolase (FAH; E.C.3.7.1.2), the last enzyme in the catabolic pathway of tyrosine. FAH was purified from rat and human liver and was used to immunize rabbits. Specific antibodies were used to probe protein extracts of livers and other tissues of normal and tyrosinemic patients. No immunoreactive FAH band was observed on immunoblots of liver, kidneys, and lymphocytes from patients presenting with the acute form of hereditary tyrosinemia. Patients with the chronic form had immunoreactive FAH at a level approximately 20% of normal liver values, which was correlated with the measured enzymatic activity. Immunoblot analysis of aborted fetal tissues revealed normal FAH immunoreactivity in normal liver and kidneys. No FAH immunoreactivity was found in liver and kidneys of tyrosinemic fetuses. The presence of FAH immunoreactivity in normal fetal tissues suggests that deficient FAH activity in tyrosinemia is not simply related to a developmentally regulated expression of the enzyme. By this immunoblot assay, FAH was detected in most human tissues, with maximal immunoreactivity in liver and kidneys and with only trace amounts in chorionic villi and cultured amniocytes. These data confirm that the primary defect in the acute form of hereditary tyrosinemia is an absence of FAH. Moreover, these data suggest that both clinical forms of the disease have a different molecular basis.

Fluorometric quantitative tyrosine determination on dried-blood spots is the primary neonatal screening test used for tyrosinemia type I (HT) in the province of Quebec. Succinylacetone determination on these same spots is used as the complementary test when the tyrosine level is higher than a given threshold. This procedure has proved to be less discriminant over the past few years because of changes in newborn feeding and because of early discharge of newborns from the nursery. We have developed an enzyme-linked immunosorbent assay (ELISA) to measure the deficient enzyme in HT in dried-blood spots. Fumarylacetoacetase (FAH) (E.C.3.7.1.2) was measured retrospectively by an ELISA on 25 dried-blood samples from proven patients with HT and prospectively in 72,000 specimens received in the neonatal screening program. In this pilot project, FAH was measured first, and, if necessary, succinylacetone was determined as the complementary test. All 25 samples from proven patients and specimens from four other patients detected in the pilot study have shown almost complete absence of FAH in dried-blood samples. At a cutoff level of 12.5% of normal adult blood spotted on the same type of paper, only 30 other cases disclosed FAH levels low enough to warrant succinylacetone measurement but had no detectable succinylacetone. The false-positive rate is thus 1:2,400 with this primary ELISA. However, blood transfusion in newborns prior to blood collection on filter paper may yield false-negative tests, since FAH is present in erythrocytes of normal donors.(ABSTRACT TRUNCATED AT 250 WORDS)

Heat-shock protein (hsp) 83 was purified from Drosophila culture cells. Analysis by gel filtration revealed that this hsp exists in a dimeric form under nondenaturing conditions. Monoclonal and polyclonal antibodies produced against this hsp have been used to determine its intracellular localization by indirect immunofluorescence and immunogold electron microscopy in normal cells, after heat shock, during recovery and after a second heat shock. Under normal conditions, hsp 83 is predominantly cytoplasmic. Immunogold labeling reveals that this hsp is associated with vacuole-like structures containing numerous dense bodies. In addition, hsp 83 is detected, albeit at a lower level, in the nucleus where it is found within the network of perichromatin ribonucleoprotein (RNP) fibrils. This distribution changes during heat shock: hsp 83 is then found in increased concentrations at the cell periphery close to the plasma membrane. After a recovery period, hsp 83 appears associated with the nuclear membrane and/or with the neighboring endoplasmic reticulum. Following a second heat shock at 37 degrees C after recovery, a renewed deposition of hsp 83 is observed at the cell periphery. A small population of cells also shows an increased concentration of this protein in the nucleus. This intracellular distribution of hsp 83 is consistent with its reported association with various cellular proteins and suggest that this hsp may be involved in their intracellular transport and/or in the modulation of their activity.

The means for analysing T-cell activities are very limited. Indeed, the numerical estimation of the changes which take place in human T lymphocytes had to rely solely on the observation of transformed cells (blasts). However, even if various mitogens appear selective for either T of B cells, we know that both types of lymphocytes are ultimately transformed into blasts. This is particularly true in studies conducted on mixtures of T and B cells where stimulated T cells can release substances which act on B cells. On the other hand, studies with purified populations of T or B cells do not express the cellular interactions that occur naturally between these two types of lymphocytes. For all these reasons, it seemed that the estimation of " quanta " of activity unique to T lymphocytes would contribute significantly to our knowledge of this line of cells. We describe a new method of detecting the in vitro stimulation of lymphocytes based on the appearance of cells having the property to cluster several layers of sheep red blood cells (SRBC) around themselves, forming multilayer rosettes (CFC). This formation is temperature-dependent and requires trypanblue-excluding, metabolically active blastoid cells. Non-separated cells as well as purified T cells stimulated with allogeneic leucocytes (MLR), specific antigens or polyclonal mitogens (PWM, ConA) gave rise to CFC. Such multilayer rosettes were not formed by separated B cells. In the MLR, CFC were detected 48 h after beginning of cultures and increased in number thereafter just like thymidine incorporation. The response to ConA and PWM was detected earlier and gave rise to two or three peaks, the first rise in the number of CFC coinciding with the peak of thymidine incorporation, but the maximal increase occurring two or three days later. Treatment of blastoid cells with a serum specific for T cells--but not with an anti-immunoglobulin serum--abolished their ability to form ordinary or multilayer rosettes. When separated, however, on a nylon wool column, CFC were more adherent than the bulk of T cells. It is suggested that CFC form a subpopulation of T cells with distinct characteristics, allowing a direct assessment of membrane changes which take place when T lymphocytes are activated in vitro.

Fibronectin, a human fibroblast surface and plasma protein, is present in cryoprecipitates of synovial fluids and/or sera from patients with rheumatoid arthritis, systemic lupus erythematosus, ankylosing spondylitis, and mixed essential cryoglobulinemia. Fibronectin was shown to be capable of influencing cryoprecipitate formation. No antibodies to fibronectin could be detected, thus ruling out the possibility that it was directly involved in the formation of cold-insoluble antigen--antibody complexes. Fibrinogen or fibrinogen degradation products were frequently present in synovial fluid cryoprecipitates but rarely in serum cryoprecipitates. Since complexes of fibronectin--fibrin--fibrinogen are known to be cold-insoluble, such interactions could occur in synovial fluids and contribute to the formation of cryoprecipitates. In serum, however, this is not likely to occur, and the mechanism by which fibronectin influences cryoprecipitate formation remains to be elucidated.

The last hours of intrafollicular maturation are important to preserve the potential of oocytes for fertilization and development. To collect mature oocytes it is consequently essential to predict accurately the time of ovulation. Therefore we have modified a regular and specific plasma LH radioimmunoassay to a convenient assay providing reliable results within four to five hours. Using this technique we have been able to detect the LH peak with a delay small enough to predict the moment of ovulation; then we have been able to observe and photograph ovulation. The antigen-antibody reaction takes place during a two hour incubation at 37 degrees C. The immune complex is precipitated by addition of a rabbit anti-gamma-globulin and incubation at 22 degrees C for 30 minutes. The variation between fast and regular assays is lower than 15%. From a study of 25 cycles in 12 animals we suggest intervals of approximately 27 hours between the beginning of the LH peak and ovulation and of about 17 hours between the end of the peak and ovulation. Intervals between the beginning of heat and ovulation appear irregular and cannot be used with certainty as a reference point to predict ovulation. Five ovulations were observed closely and in the five cases, the formation of an apex was noted Our results clearly show that with the technique mentioned above we can predict rapidly and precisely bovine ovulation.

The differentiation of human B lymphocytes in vitro into Ig secreting cells, as measured by a radioimmunoassay rendered specific to IgG and IgM, in response to a polyclonal stimulant, requires the participation of several cellular types. Upon poleweed-mitogen stimulation, B-cell enriched and monocyte containing cell suspensions prepared by elution of nylon wool adherent cells, when mixed with various concentrations of T cells purified by filtration through nylon wool column and depletion of EAC-rosette forming cells, synthesize 5 to 20 times more Ig than non separated lymphocytes. These experiments which demonstrate the cooperation of certain T cells also suggest that other cells eliminated by nylon wool adherence act as suppressor cells. The cooperation of these T cells is independent of their proliferation and can be obtained with allogeneic T cell.

We describe a two-site immunoradiometric assay for human alpha1-fetoprotein, with use of antibody-coated polystyrene tubes as solid phase. The sensitivity, precision, and simplicity of this system make it eminently suitable for mass-screening purposes. We currently use it for neonatal detection of hereditary tyrosinemia in the Province of Quebec; measurements are made on blood spotted and dried on paper. This system could be well suited for other mass surveys, such as prenatal screenings for fetal abnormalities.

As a step towards the cloning of the gene for fumarylacetoacetate hydrolase (FAH), we have purified the FAH mRNA from rat liver by specific immunoadsorption of polysomes. The relative abundance of this mRNA has been estimated to be 0.14%. The major in vitro translation product of the purified mRNA preparation is specifically precipitated by a rabbit anti-rat FAH antiserum and it is, furthermore, undistinguishable by criteria of mass and charge from purified rat FAH.

Hepatocytes were isolated from 1-day and 1,2,3 and 12-week old rat livers by collagenase perfusion and the relative numbers of albumin (ALB) and alpha-fetoprotein (AFP) producers were evaluated using the reverse hemolytic plaque assay. The percentage of ALB producers remained essentially constant to 35% over the 12-week period. In contrast, the percentage of AFP producers varied from 19% at 1 day up to 28% at 1 week and then down to 0.1% at 3 weeks. Moreover, a double identification of secreting hepatocytes, using an adaptation of the plaque assay, demonstrated that AFP producing hepatocytes were also ALB producers. These results are explained in terms of a restricted specialization of differentiating hepatocytes during normal development.

A total of 344 sera positive for hepatitis B surface antigen from volunteer blood donors at several Canadian Red Cross centres were subtyped for ad and ay specificity by counterelectrophoresis. Of the 50 sera from Toronto 21 (42%) were ad and 29 (58%) were ay; of the 95 from Montreal 82 (86%) were ad and 13 (14%) were ay; of the 199 from Quebec 179 (90%) were ad and only 20 (10%) were ay. The w and r specificities were also determined in 125 of the samples: 123 were w; the 2 samples of r specificity were from Toronto. On the other hand, among 45 sera from patients with acute hepatitis type B in Quebec 13 (29%) were ad and 33 (71%) ay.

We have recently reported that fibronectin is a component of mixed cryoglobulins and is responsible in large part for cryoprecipitate formation. We have now extended our studies to serum cryoprecipitates formed in the presence of monoclonal cryoglobulins in patients with lymphoproliferative diseases in order to investigate for a possibly similar influence of fibronectin on cryoprecipitation. Results have shown that these cryoprecipitates contain fibronectin in proportions varying between 10% and 14% of their total protein content. Furthermore, SDS-polyacrylamide gel electrophoresis revealed that next to the cryoglobulins, fibronectin constitutes the second major component of the cryoprecipitates. Depletion of the sera in fibronectin prior to exposure to 4 degrees C, markedly reduced cryoprecipitation of the cryoglobulins, whereas reconstitution in fibronectin restored it. These studies support and extend our previous findings showing the important influence exerted by fibronectin on the process of immunoglobulin cryoprecipitation.

Radioallergosorbent test (RAST) for the measurement of IgE antibodies has been introduced more than 15 years ago and a number of technical modifications have since improved its sensitivity and reproducibility. The test has been applied to the diagnosis of allergy and to determine changes in the levels of IgE antibodies following immunotherapy. However, specific IgG antibodies are raised during such a therapy and can interfere with the RAST. We have developed a reverse enzymoallergosorbent test (REAST) where microtiter plates are first coated with a purified polyclonal anti-IgE antibody, then with the serum to test and finally with peroxidase-labeled antigen. This assay is antigen specific as shown by the significant inhibition of binding of the labeled antigen in presence of unlabeled specific antigen (greater than 95%) and the absence of inhibition in presence of irrelevant antigens. The values found in atopic patients (85 subjects) were significantly higher than in the non-atopic donors (35 subjects) (1.14 U +/- 1.20 vs. 0.01 U +/- 0.02, P less than 0.0005) and there was a good correlation with the Pharmacia RAST (P less than 0.0005). The levels of specific IgE by both REAST and RAST correlated well with the clinical symptomatology.

Monitoring of donors with selective IgA deficiency, i.e., with less than 0.5 mg IgA/l, led to the observation that serum IgA levels wer not constant. Small but significant fluctuations in IgA levels were noted (coefficient of variance: 143%) which were greater than the variability inherent in the testing methodology (coefficient of variance: 10%). These fluctuations created difficulties in terms of defining IgA-deficient blood products and had implications with respect to the mechanisms involved in IgA deficiency. With respect to supplying IgA-deficient blood products, in our experience a cutoff level of 0.5 mg/l should be the maximum permissible IgA concentration in order to ensure that no adverse reactions occur in individuals with class-specific anti-IgA.

A commonly used and specific plasma LH radioimmunoassay was modified to provide reliable results for cow LH concentrations in blood within 4-5 h. Blood samples were collected at 2- or 4-h intervals from at least 24 h before the expected oestrus until after its onset. Thereafter, samples were collected at intervals of 2 or 4 days until the next oestrus. For the assay, the antigen-antibody reaction took place during a 2-h incubation at 37 degrees C. The immune complex was precipitated by addition of rabbit anti-gamma-globulin and polyethylene glycol followed by an incubation at 22 degrees C for 30 min. The coefficient of variation between this assay and the standard assay was less than 15%. From a study of 25 cycles from 12 animals, we obtained intervals of congruent to 27.3 h between the beginning of the LH peak and ovulation and congruent to 17.5 h between the end of the peak and ovulation. This modified technique can be used to predict rapidly and precisely the time of ovulation in cattle.

This paper describes a modification of the reverse hemolytic plaque assay allowing the enumeration of immunoglobulin (Ig)-secreting lymphocytes at rest or after in vitro stimulation. Ig-secreting cells are mixed with anti-Ig coated sheep red blood cells, developing anti-Ig antiserum and complement in liquid medium. This mixture is pipetted between two slides and the hemolytic plaques are numerated after incubation at 37 degrees C. The class of Ig secreted can be determined using a monospecific developing antiserum. This technique is easier to perform and more economical than the previously described method using agarose without loss of sensitivity, the number of plaques detected being about the same and the kinetic of their appearance comparable.

The two-step collagenase perfusion method originally developed for the high yield isolation of parenchymal cells from adult rat livers has been adapted to rats of 1 day, 1 week, and 2 weeks of age. The use of this method to isolate hepatocytes from five or six rats of the respective ages demonstrated its reliability in terms of cell yield, percentage of single cells, and cell viability. In all cases, hepatocytes attach with high efficiency to fibronectin precoated dishes using serum-free culture medium. The dynamics of spreading is faster for newborn hepatocytes than adult ones. The functional integrity of these parenchymal liver cells was assessed by their capacity to secrete albumin and alpha-fetoprotein in serum-free medium and to express lactate dehydrogenase activity over a 24-hr period in primary culture.

Liver cells were isolated by collagenase perfusion from rats of 1 day, and 1, 3, 5, and 12 weeks of age, fractionated by velocity sedimentation at 1 g (STAPUT), and the major cell types were identified in terms of specific functions. Alphafetoprotein and albumin were used as markers of differentiating hepatocytes and these functional activities were evaluated in a quantitative manner using a radioimmunoassay. The capacity of this cell type to store 35S-BSP, an indicator of bile formation, was also evaluated. Sinusoidal cells and hematopoietic cells were identified on the basis of their ability to take up 99mTC-colloid sulfur and to incorporate 59Fe, respectively. The fractionation procedure allowed a good separation of sinusoidal cells from hepatocytes at all postnatal ages and also of erythroid cells still present during the first week after birth. With increasing age, alphafetoprotein-producing hepatocytes exhibited changes in sedimentation velocities that parallelled those of albumin-producers. In turn, the latter hepatocyte subpopulation underwent gradual shifts in modal peak velocities similar to those of bile-forming hepatocytes. The fractionated hepatocytes obtained at different ages were further analyzed in terms of cell volume and nuclear ploidy using a Coulter counter system. This quantitative analysis obtained at the cellular level demonstrated that during the age-related differentiation of hepatocytes, which occurs during the postnatal period and results in the gradual appearance of cells of higher ploidy levels, the extent of albumin production and bile formation can be correlated with the hepatocyte volume.

To gain insight on the possible functions of heat shock proteins (hsp's) in Drosophila, we have purified the 83-kilodalton hsp (hsp 83) from cultured cells and studied its intracellular localization by immunofluorescence in normal, heat-shocked, and recovering cells. The specificity of the antibody was assessed by one- and two-dimensional gel immunoblotting and by partial proteolytic digestion. The anti-hsp 83 antibody does not show any significant cross-reactivity with hsp's of different avian or mammalian cell lines, but cross-reacts with hsp's of similar molecular masses in other dipteran insects. The partial proteolytic peptide maps of Drosophila hsp 83 differ from those of mouse hsp 89 and chicken hsp 84. Immunoblotting of Drosophila Kc cells heat shocked at different temperatures indicates a maximal expression of hsp 83 at 33 degrees C. By immunofluorescence, hsp 83 is shown to have a strictly cytoplasmic localization. In unstressed cells, it is distributed in the entire cytoplasm with a slight enrichment in the perinuclear region. After heat shock, it seems to concentrate at the cell periphery close to the plasma membrane and it gradually redistributes to the whole cytoplasm during cellular recovery at normal temperatures.

Eighty-five cases of beta-thalassemia minor were found between January 1975 and November 1977 in 18 families of French-speaking Quebeckers without Mediterranean ancestry. Most of the families had settled in Quebec more than 200 years ago, largely in Portneuf county. This is the first report of such a number of cases of beta-thalassemia in this population. Thus, beta-thalassemia minor is a relatively common condition in Quebec and must be considered in cases of microcytic anemia without evidence of iron deficiency. The hematologic findings were similar to those reported in the past in other populations. Two cases of delta beta-thalassemia minor (in sisters) are also reported; this is the first report of such cases in French Canadians.

Albumin-secretins hepatocytes were detected as plaque-forming cells when incubated in a complete culture medium containing sheep eythrocytes coated with purified anti-albumin, plus anti-albumin serum and complement added at appropriate dilutions. Plaque-formation is observed after incubation at 37 degrees C for 1 h 1/2 between two properly spaced microscope slides sealed with paraffin. Plaque enumeration revealed that 17-18% of normal adult rat lover cells secreted albumin. The assay which is reproductible and highly sensitive is also done in a single step.

Using monoclonal antibodies the immunogenic and allergenic characteristics of rye Group I were redefined by SDS-PAGE analysis and immunoblotting. The purified rye Group I from NIH possesses a major component of approximately 34000 Da against which most of the sera from grass-sensitive patients and none from non-atopic volunteers contain specific IgE antibodies. Monoclonal antibodies to rye Group I were raised and used to purify the antigen and to verify the cross-reactivity between grass extracts. The 3 monoclonal antibodies studied recognized different components of timothy grass and 2 of them recognized kentucky june grass but none recognized components of ragweed extracts.

Blood-group-specific A1, B, AB, H and Lea neutral glycoproteins were isolated from suitable pools of human normal meconiums by a preliminary fractionation with a cationic detergent at pH5 and 9 (borate), followed by ion-exchange and gel chromatography. The ABH materials have sedimentation coefficients of about 10S-11S, whereas the Lea preparation, not strictly homogeneous, shows a coefficient of 7S. From the detailed analytical data collected, the following relations are deduced between these various substances; they all possess a common peptide core; there are stable ratios of N-acetylglucosamine/N-acetylgalactosamine in the B, H and Lea materials and of N-acetylglucosamine/galactose in A, H and Lea materials, from which the numbers of A and B determinants are estimated. In the ABH substances, the ratio of glucosamine to the sum of threonine and serine is stable. Presumably because of genetic factors, the amount of fucose varies among the different glycoproteins, but it is always definitely lower than in the average cyst substances. Various serological tests and precipitin methods were used to measure the potency, purity and integrity of the preparations, including comparisons between A1 and A2 substances from this source. The Leb activity did not appear as high as it is in glycoproteins from adults and a possible interpretation would be the immature Lewis system as observed on erythrocytes; this could explain their very strong inhibiting power towards iso-agglutinins. This family of substances with various specificities has common features with that prepared from ovarian cysts, but differs clearly on some points.