Development of Lactococcus lactis encoding fluorescent proteins, GFP, mCherry and iRFP regulated by the nisin-controlled gene expression system.

Fluorescent proteins are useful reporter molecules for a variety of biological systems. We present an alternative strategy for cloning reporter genes that are regulated by the nisin-controlled gene expression (NICE) system. Lactoccocus lactis was genetically engineered to express green fluorescent protein (GFP), mCherry or near-infrared fluorescent protein (iRFP). The reporter gene sequences were optimized to be expressed by L. lactis using inducible promoter pNis within the pNZ8048 vector. Expression of constructions that carry mCherry or GFP was observed by fluorescence microscopy 2 h after induction with nisin. Expression of iRFP was evaluated at 700 nm using an infrared scanner; cultures induced for 6 h showed greater iRFP expression than non-induced cultures or those expressing GFP. We demonstrated that L. lactis can express efficiently GFP, mCherry and iRFP fluorescent proteins using an inducible expression system. These strains will be useful for live cell imaging studies in vitro or for imaging studies in vivo in the case of iRFP.

Ciliate species diversity and host-parasitoid codiversification in Pseudocollinia infecting krill, with description of Pseudocollinia similis sp. nov.

All parasitoid apostome ciliates infecting krill in the northeastern Pacific are currently assigned to the genus Pseudocollinia. Each krill specimen is apparently infected by only 1 Pseudocollinia species. We describe Pseudocollinia similis sp. nov., discovered infecting the krill Thysanoessa spinifera off Oregon, USA. Its protomite-tomite stage resembles that of P. beringensis, which infects T. inermis (type host species), T. longipes, and T. raschii females in the Bering Sea. These ciliates have similar numbers of somatic kineties (18-21 vs. 16-20) and typically have 3 oral kineties. Furthermore, these 2 apostomes are sister species on gene trees based on sequences of small subunit rRNA (0.06% difference) and cytochrome c oxidase subunit 1 (cox1; 30% difference). P. brintoni and P. oregonensis are closely related as a separate group from P. similis and P. beringensis. The similar tree topologies based on the cox1 sequences of 21 host krill individuals representing 6 krill species (Euphausia pacifica, Nyctiphanes simplex, T. inermis, T. longipes, T. raschii, and T. spinifera) and the apostomes isolated from these krill suggest host-parasitoid codiversification. However, this hypothesis was statistically rejected by an approximately unbiased test in which the host tree topology was used to model parasitoid evolution (p ≤ 0.05).

Pseudocollinia brintoni gen. nov., sp. nov. (Apostomatida: Colliniidae), a parasitoid ciliate infecting the euphausiid Nyctiphanes simplex.

A novel parasitoid ciliate, Pseudocollinia brintoni gen. nov., sp. nov. was discovered infecting the subtropical sac-spawning euphausiid Nyctiphanes simplex off both coasts of the Baja California peninsula, Mexico. We used microscopic, and genetic information to describe this species throughout most of its life cycle. Pseudocollinia is distinguished from other Colliniidae genera because it exclusively infects euphausiids, has a polymorphic life cycle, and has a small cone-shaped oral cavity whose left wall has a field of ciliated kinetosomes and whose opening is surrounded on the left and right by 2 'oral' kineties (or ciliary rows) that terminate at its anterior border. Two related species that infect different euphausiid species from higher latitudes in the northeastern Pacific Ocean, Collinia beringensis Capriulo and Small, 1986, briefly redescribed herein, and Collinia oregonensis Gómez-Gutiérrez, Peterson, and Morado, 2006, are transferred to the genus Pseudocollinia. P. brintoni has between 12 and 18 somatic kineties, and its oral cavity has only 2 oral kineties, while P. beringensis comb. nov. has more somatic kineties, including 3 oral kineties. P. oregonensis comb. nov. has an intermediate number of somatic kineties. P. beringensis comb. nov. also infects Thysanoessa raschi (a new host species). SSU rRNA and cox1 gene sequences demonstrated that Pseudocollinia ciliates are apostome ciliates and that P. brintoni is different from P. beringensis comb. nov. High densities of rod-shaped bacteria (1.7 µm length, 0.2 to 0.5 µm diameter) were associated with P. brintoni. After euphausiid rupture, high concentrations of P. brintoni and bacteria cluster to form 3 to 6 cm long filaments where tomites encyst and transform to the phoront stage; this is a novel place for encystation. P. brintoni may complete its life cycle when the euphausiids feed on these filaments.

Utilidad de la fibrobroncoscopia en las intubaciones traqueales difíciles / Usefulness of fiberoptic bronchoscopy for difficult tracheal intubations

La fibrobroncoscopia (FB) representa una gran ayuda para la realización de intubaciones difíciles o fallidas por los métodos convencionales. Nuestro objetivo fue conocer el número de intubaciones realizadas mediante FB, así como características de las mismas y analizar los problemas detectados en base a la experiencia acumulada. Metodología: Se recogieron todas las intubaciones realizadas con control de FB, desde enero de 1996 hasta Octubre de 2005.En ésta figuraban el servicio solicitante de la intubación y el de procedencia del paciente, el motivo de la intubación y complicaciones de la técnica. Resultados: Se realizaron un total de 4291 FB delas cuales el 11% fueron intubaciones guiadas con FB. La edad media fue 53,6 años, siendo el 77,2% varones. Casi todas las intubaciones fueron solicitadas por el servicio de anestesia (95,7%).Los servicios de procedencia de los pacientes fueron cirugía general32%, cirugía maxilofacial (MXF) 31,1%, traumatología 14,7%,Otorrinolaringología (ORL) 8,3%, Unidad de Cuidados Intensivos/Unidad de Reanimación Postoperatoria 4,3%, neurocirugía4,1% y otros 5,3%. Las indicaciones fueron intubaciones bronquiales selectivas 23%, tumores MXF 22,6%, rigidez cervical8,5%, limitación en la apertura de la boca 8,3%, obesidad 7%,tumores ORL 4,3%, fracturas MXF, síndrome de apnea del sueño y espondilitis anquilosante el 3,6% respectivamente, patología tiroidea, artritis reumatoide y reintubación el 3,2% y otros en el5,3%. Se produjeron complicaciones (4,2%) todas de escasa importancia. En 5 casos (1%) no se consiguió la intubación con control FB. Conclusiones: la intubación guiada con FB es una técnica segura y con una elevada rentabilidad en los casos de intubaciones difíciles o fallidas con los medios convencionales

The fiber optic bronchoscopy (FB) represents a great help for difficult or failed intubations by conventional methods. The objective was to know the number of intubations performed by FB, as well as the characteristics of these intubations and, in our experience, to analyze the problems detected during these intubations. Methodology: we analyzed in a data base all the intubations performed by FB guidance, from January 1996 to October 2005. In this database we recorded the services that had asked for the intubation, the admission service of the patient, the reason for the intubation and complications. Results: From a total of 4.291 FB performed,11% were FB to guide intubations. Almost all the intubations were asked for by the an a esthesiology service (95.7%).The admission services of the patients were General Surgery 32%,Maxillofacial (MF) Surgery 31.1%, Traumatology 14.7%, Otorhinolaryngology(ORL) 8.3%, Intensive Care Unit/Postoperative Reanimation Unit 4.3%, Neurosurgery 4.1%, and other services5.3%. The most frequent reasons for the intubations were selective bronchial intubations 23%, MF tumors 22.6%, cervical rigidity8.5%, limitations in the opening of the mouth 8.3%, obesity 7%,ORL tumors 4.3%, MF fractures, sleep apnea syndrome and ankylosing spondylitis 3.6% respectively, thyroid diseases, rheumatoid arthritis and reintubation 3.2% and other 5.3%. We recorded a4.2% rate of mild complications, all of little importance. In 5 cases(1%) the intubation guided by FB was impossible. Conclusions: The intubation guided by FB is a safe technique and with a high yield in the cases of difficult or failed intubations by conventional methods

[Comparison of two models of education for asthmatic patients]. / Comparación de dos modelos de educación para pacientes asmáticos.

BACKGROUND: The education programs have demonstrated to be an important point in the management of asthmatic patients. The aim of the present study was to assess if an intensive group asthma education program was able to improve a simplified and individual asthma education program, both with a self-management plan included. PATIENTS AND METHOD: A prospective randomised controlled trial was conducted over 12 months and 73 moderate-severe asthmatic patients were included. Patients were randomly assigned to control or study group. Patients in control group received individual and simplified education with a self-management plan and patients in study group attended an <> in small groups where they received a three hours education program in three weeks.The outcome measures were: number of hospital admissions, observation unit admissions, emergency visits and unscheduled general practice consultations. Knowledge, adherence to treatment, ability with inhaler devices, airway function, environmental control measures and quality of life. They were assessed at the beginning and the end of the study. RESULTS: At the end of the study period the outcome of morbidity had improved in both groups but the number of observation unit admissions (p = 0.028) and unscheduled general practice consultations (p = 0.022) was substantially lower in study group. This group also demonstrated improvement in environmental control measures and better inhaler technique and knowledge (p = 0.007). In adherence, lung function or quality of life there were no differences between groups. CONCLUSIONS: In patients with asthma better results can be obtained with an intensive group asthma education program than with the individual and simplified program.

Urea equilibrium unfolding of the major core protein of the retrovirus feline immunodeficiency virus and its tryptophan mutants.

Circular dichroism and fluorescence spectroscopy have been employed to study the urea unfolding mechanism of a recombinant form of the major core protein of feline immunodeficiency virus (FIV-rp24) and its native tryptophan mutants. The equilibrium denaturation curves indicate the existence of two transitions. The first unfolding transition most likely reflects the denaturation of the carboxy-terminal region of FIV-rp24. Consequently, the second transition, where the changes in fluorescence are produced, should reflect the denaturation of the amino-terminal region. If the intermediate observed upon urea denaturation is an on-pathway species, the data described herein can reflect the sequential and independent loss of structure of the two domains that this type of proteins possesses.

Cloning, expression, and purification of histidine-tagged preS domains of hepatitis B virus.

The preS domains of the hepatitis B virus are hydrophilic polypeptides that have been implicated, among other functions, in the binding of the virus to hepatocytes and in the induction of virus-neutralizing antibodies. A method of overproducing the preS domains of two different subtypes, adw and ayw, has been developed by adding a 6x His tag at the carboxy-terminal end of the polypeptides. Codons for the 6x His were added in reverse primers used to amplify the corresponding cDNAs. The polymerase chain reaction products were cloned into the expression vectors pET-3d (subtype ayw) and pT7-7 (subtype adw), under the control of the inducible bacteriophage T7 RNA polymerase promoter. Upon induction with isopropyl-beta-d-thiogalactopyranoside, proteins were overexpressed and purified by affinity chromatography on a Ni-nitrilotriacetic acid agarose column. This method yielded 20-40 mg of highly pure and very stable proteins per liter of cell culture. Circular dichroism and fluorescence spectroscopy of isolated preS-his-ayw and preS-his-adw, as well as their ability to bind polymerized human serum albumin, indicate that the 6x His tag does not modify the native-like conformation and, therefore, they may be considered as useful tools to study the function of these viral polypeptide regions.

Community structure of zooplankton in the main entrance of Bahía Magdalena, México during 1996.

The zooplankton community structure, including copepods, euphausiids, chaetognaths, and decapod larvae, was monitored during six circadian cycles using Bongo net (500 microns mesh net) samples from Bahía Magdalena, on the southwest coast of Baja California, México. Samples were obtained during three oceanographic surveys (March, July, and December 1996) to describe the changes in the zooplankton community structure throughout the main mouth of Bahía Magdalena. The zooplankton community structure showed strong changes with a close relation to environmental conditions. During March, a well-mixed water column with low temperature and salinity indicated an influence of the California Current water and local upwelling processes. During July, temperature increased and a wide salinity range was recorded. The stratification of the water column was intense during summer, enhancing the thermocline. The highest temperatures and salinity were recorded in December, related to the presence of the Costa Rica Coastal Current (CRCC). The thermocline deepened as water temperature increased. A typical temperate community structure with low specific richness dominated by Calanus pacificus, Nyctiphanes simplex, and Acartia clausi and high zooplankton biomass (average 9.3 and 5.5 ml 1000 m-3 respectively) during March and July shifted to a more complex tropical community structure with a low zooplankton biomass in December (average 0.37 ml 1000 m-3). The mouth of Bahía Magdalena has a vigorous exchange of water caused by tidal currents. The zooplankton community structure was not significantly different between the central part of Bahía Magdalena and the continental shelf outside the bay for all months. The results suggest a more dynamic inside-outside interaction of zooplankton assemblages than first thought.

Circular dichroism and fluorescence spectroscopic properties of the major core protein of feline immunodeficiency virus and its tryptophan mutants. Assignment of the individual contribution of the aromatic sidechains.

The gene coding for the major capsid protein of feline immunodeficiency virus (FIV) has been cloned into the expression vector pQE60, which allows protein purification by affinity chromatography on a nitrilotriacetic acid/Ni/agarose column. The gene was expressed in Escherichia coli and the resultant soluble protein (FIV-rp24) purified to electrophoretic homogeneity. The amino-acid composition of the recombinant protein is almost identical to that predicted from the DNA sequence. This protein has two tryptophan residues at positions 40 and 126 that have been replaced by phenylalanine by site-directed mutagenesis to obtain two single mutants and a double mutant. Circular dichroism and fluorescence spectroscopy were employed to study the structural features of FIV-rp24 protein and its tryptophan mutants. The analysis of the CD spectra indicated that alpha-helix is the major secondary structural element (48-52%) and that the overall three-dimensional structure is not modified by the mutations. The fluorescence emission spectra showed that both tryptophan residues occupy a highly hydrophobic environment. Moreover, the different tyrosine fluorescence intensities of wild-type and mutant proteins are indicative of the existence of resonance energy transfer processes to nearby tryptophan. The individual contributions of each tryptophan residue to the spectroscopic properties of the wild-type protein were obtained from the spectra of all these proteins. Thermal denaturation studies indicate that the two tryptophan residues do not contribute equally to the stabilization of the three-dimensional structure.

Fusogenic activity of hepadnavirus peptides corresponding to sequences downstream of the putative cleavage site.

Sequence homology between the amino-terminal region of the S protein of hepatitis B Virus (HBV) and known fusion peptides from retroviruses and paramyxoviruses led us to propose that this region might be equally involved in the initial infective steps of hepadnaviruses. In fact, we showed that a synthetic peptide corresponding to the N-terminus region of the S protein of HBV had membrane-interacting properties and was able to induce liposome fusion adopting an extended (beta-sheet) conformation (Rodríguez-Crespo et al., 1996, 1995). We describe herein studies on the interaction of peptides derived from the N-terminal region of the S protein of duck (DHBV: Met-Ser-Gly-Thr-Phe-Gly-Gly-Ile-Leu-Ala-Gly-Leu-Ile-Gly-Leu-Leu) and woodchuck hepatitis B viruses (WHV: Met-Ser-Pro-Ser-Ser-Leu-Leu-Gly-Leu-Leu-Ala-Gly-Leu-Gln-Val-Val) with liposomes. These peptides were able to induce to a different extent aggregation, lipid mixing, and leakage of internal aqueous contents from both neutral and negatively charged phospholipid vesicles in a concentration-dependent and pH-independent manner. Fluorescence depolarization of 1,6-diphenyl-1,3,5-hexatriene-labeled vesicles indicated that both peptides become inserted into the hydrophobic core of the lipid bilayer. Circular dichroism studies indicated that the DHBV peptide adopts an extended conformation in the presence of lipids, whereas the WHV peptide displays a high content of alpha-helical conformation. Therefore, these results extend our previous findings obtained for human hepatitis B virus to other members of the hepadnavirus family and suggest that this region of the S protein is important in the initial steps of the infective cycle.

Structural properties of the putative fusion peptide of hepatitis B virus upon interaction with phospholipids. Circular dichroism and Fourier-transform infrared spectroscopy studies.

A peptide corresponding to the N-terminal sequence of the S protein from hepatitis B virus (Met-Glu-Asn-Ile-Thr-Ser-Gly-Phe-Leu-Gly-Pro-Leu-Leu-Val-Leu-Gln) has been previously shown to interact with phospholipids and promote vesicle aggregation, phospholipid mixing, and liposome leakage, as well as erythrocyte lysis [Rodríguez-Crespo, I., Núñez, E., Gómez-Gutiérrez, J., Yélamos, B., Albar, J. P., Peterson, D. L. & Gavilanes, F. (1995) J. Gen. Virol. 76, 301-308]. The conformation of this putative fusion peptide has been studied, both at low and high peptide concentrations, by means of circular dichroism and Fourier-transform infrared spectroscopy, respectively. When the peptide is dissolved in trifluoroethanol, a significant population of alpha-helical structure is found in spite of the proline residue at position 11. In contrast, this hydrophobic oligopeptide has a high tendency to form large beta-sheet aggregates in aqueous buffers. Most of these aggregates can be eliminated by centrifugation. The peptide remaining in the supernatant adopts a non-ordered conformation. The aggregates can be dissociated by the anionic detergent sodium cholate, but the peptide still maintains an extended conformation. In the presence of acidic phospholipid vesicles, the putative fusion peptide adopts a highly stable beta-sheet conformation. Thus, unlike the fusion peptides of other viruses, an extended conformation seems to be the preferred structure when interacting with phospholipids. Such a conformation should be responsible for its membrane destabilization properties.

Oculophryxus bicaulis, a new genus and species of dajid isopod parasitic on the euphausiid Stylocheiron affine Hansen.

A new genus and species of dajid isopod is described from the euphausiid Stylocheiron affine Hansen. The isopod is a member of the Dajidae Krøyer, 1842, which are ectoparasites of shrimp, mysids, and krill. The female of the new genus and species is unique in its attachment to the eyestalk of its euphausiid host. The well-developed antennae encircle the eye peduncles of the host. The parasite presumably feeds by sucking blood directly from the head of the euphausiid. The new genus can be distinguished from other genera by the elongate, spoon-shaped antennae, the number of pereopods, and the indistinct abdomen. The male parasite attaches to the posterior of the female near the margin of the marsupium. The male can be distinguished from other genera by the rudimentary 7th pereopod. Epicaridia, microniscus and cryptoniscus larvae were not observed. The parasite was found in samples taken during 7 oceanographic surveys made along the west coast of Baja California. The parasite was found on the furciliae, juveniles, and immature males and females of S. affine. Female isopods may castrate their hosts as none of the infected hosts (n = 27) had reached maturity. Of the 3 eco-phenotypes of S. affine that are found in the region, the isopod apparently prefers the California Current morph. However, in the Gulf of Mexico, a similar parasite was found on S. longicorne Sars.

Antigenicity of hepatitis B surface antigen proteins reconstituted with phospholipids.

Hepatitis B surface antigen (HBsAg) has been reconstituted with different phospholipid classes. All epitopes defined by a panel of monoclonal antibodies which recognize both group- and subtype-specific antigenic determinants showed specificity for acidic phospholipids. Electrostatic interactions between HBsAg proteins and acidic phospholipids are partly responsible for the complete recovery of the antigenic properties. In addition to the nature of the polar head group, the fatty acid composition of the phospholipid also influenced the recovery of the antigenic activity. Negatively charged phospholipids must bear at least one unsaturated fatty acid in order to be effective in recovering full antigenic activity of HBsAg. The results reported herein support the conclusion that the antigenic activity is dependent on the physical state of the phospholipid moiety. The appropriate membrane fluidity is required for optimum conformation but, once this conformation is established, additional interactions imparted by the various phospholipids give a difference in the patterns of antigenicity. The analysis of binding of the monoclonal antibodies allowed the classification of the epitopes into two groups according to their dependence on the lipid moiety. Of all the antigenic determinants only those close to the lipid-protein interface would change upon direct interaction with the phospholipids. The rest would depend on the correct protein conformation determined by the appropriate phospholipid composition.

Phospholipid interactions of the putative fusion peptide of hepatitis B virus surface antigen S protein.

One of the first steps in the infective cycle of an enveloped virus consists of the fusion of the viral and cellular membranes. This process is usually achieved as a result of membrane destabilization brought about by a viral fusion peptide located at the amino terminus of one of the viral envelope glycoproteins. Previous sequence similarity studies by Rodríguez-Crespo et al. (Journal of General Virology 75, 637-639, 1994) have shown that a hydrophobic stretch in the amino-terminal sequence of the S protein of hepatitis B virus shares several characteristics with fusion peptides of retroviruses and paramyxoviruses. A 16 residue peptide with this sequence was synthesized and its interaction with liposomes characterized. This peptide was able to mediate vesicle aggregation, lipid mixing and liposome leakage in a pH dependent manner at concentrations ranging from 3.5 to 52.0 microM. These effects were specific for negatively charged phospholipid vesicles. The peptide was also able to haemolyse erythrocytes. This study supports the notion that the sequence might be important in the initial infective steps of this virus, interacting with the target membranes and bringing about their subsequent destabilization.

Reconstitution of hepatitis B surface antigen proteins into phospholipid vesicles.

Hepatitis B surface antigen (HBsAg), devoid of 75% of its total lipids has been reconstituted with several phospholipids by the detergent dialysis method, using the non-ionic detergent beta-D-octyl glucoside. Upon reconstitution with both neutral and acidic phospholipids, HBsAg particles had the same morphology and, as indicated by trypsin hydrolysis, the topology of the surface proteins was maintained. However, only negatively charged phospholipids were able to completely revert the conformational changes which had been induced by removal of the lipids. The helical content, as indicated by CD techniques, and the antigenic activity, as measured by binding to polyclonal antibodies, of HBsAg reconstituted with acidic phospholipids were practically identical to those of the native antigen. Cholesterol had no effect on the antigenic activity recovered by reconstitution with any of the phospholipids.

Site-directed mutagenesis of cysteine residues of hepatitis B surface antigen. Analysis of two single mutants and the double mutant.

The structure of hepatitis B surface antigen (HBsAg) is mainly maintained by an intricate disulfide network responsible for most of its structural and antigenic properties. Characterization of three cysteine-replacement mutants of HBsAg has been performed by both structural and immunological methods. Replacement of Cys121 or Cys124 with serine results in mutant proteins that show diminished binding titres to both monoclonal antibodies and to a polyclonal serum, indicating that a structural change has taken place. Circular dichroism analysis shows that the substitution of either of these two residues also diminishes the helical content of the protein. However, the double mutant, in which both cysteine residues have been simultaneously changed, reverts the properties of the single mutations, and shows similar behaviour to the wild-type protein. Both the single and double cysteine mutants are efficiently glycosylated and secreted from Chinese hamster ovary cells and, in all cases, the mutant proteins assemble into spherical particles of similar buoyant density to both the wild-type and serum derived HBsAg.

Prediction of a putative fusion peptide in the S protein of hepatitis B virus.

Sequence analysis of the S protein of hepatitis B virus (HBV) reveals a stretch of 23 hydrophobic amino acids in the amino-terminal region which shows a high degree of similarity with known fusogenic peptides from other viruses. Additionally, this sequence also appears to be highly conserved within the hepadnavirus family. Taken together, the different criteria used in this work suggest fusogenic activity in the amino-terminal region of the S protein of the envelope of HBV.

Thermal stability of hepatitis B surface antigen S proteins.

Thermal stability of hepatitis B surface antigen (HBsAg) has been studied by analyzing alterations in the native secondary structure and the antigenic activity. After heating for 19 h, circular dichrosim showed a cooperative transition with a midpoint at 49 degrees C. The conformational changes induced by temperature reduced the helical content of HBsAg S proteins from 49% at 23 degrees C to 26% at 60 degrees C and abolished the antigenic activity, as measured by binding to polyclonal antibodies. Furthermore, the six different antigenic determinants recognized by our panel of monoclonal antibodies were also shown to be dependent on the native structure of HBsAg proteins. Hence, it can be inferred that these epitopes are conformation-dependent. Binding of monoclonal antibodies to HBsAg protected the native structure of the corresponding antigenic determinant from thermal denaturation. In fact, binding of one of the monoclonals tested resulted not only in protection of the corresponding epitope, but also in a consistent increase of antibody binding with increasing temperature. Such an increase in antibody binding occurred simultaneously with an increase in the fluidity of surface lipid regions, as monitored by fluorescence depolarization of 1-(trimethylammoniophenyl)-6-phenyl-1,3,5-hexatriene. This correlation, along with the observation that lipids play an important role in maintaining the structure and antigenic activity of HBsAg (Gavilanes et al. (1990) Biochem. J. 265, 857-864), allow to speculate the certain epitopes of HBsAg which are close to the lipid-protein interface, are dependent on the fluidity of the surface lipid regions. Thus, any change in the physical state of the lipids could confer a different degree of exposure to the antigenic determinants.

Hepatitis B surface antigen. Role of lipids in maintaining the structural and antigenic properties of protein components.

Most of the lipid components of hepatitis B surface antigen (HBsAg) can be removed by treatment with the non-ionic non-denaturing detergent beta-D-octyl glucoside (OG) followed by centrifugation through caesium chloride linear density gradients (density 1.15-1.32 g/ml). The conformational changes induced by the elimination of lipids decreased the helical content of HBsAg proteins from 52 to 28% as indicated by c.d. techniques. Measurements of the extent of quenching of protein fluorescence by iodide showed that half of the tryptophan residues which are buried in the native structure of HBsAg particles are brought close to the surface of the molecule by such conformational changes. The antigenic activity, as measured by binding to polyclonal antibodies, was decreased upon removal of lipids. Moreover, the six different antigenic sites recognized by our panel of monoclonal antibodies decreased their capacity to bind to the corresponding antibody when lipids were removed. However, the extent of this decrease differed for the different antibodies. Thus the apparent dependence of antibody binding on the lipid content seemed to indicate a greater involvement of the lipid-protein interaction for some of the epitopes than for others.