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PubMed2001

Perforated microelectrode arrays implanted in the regenerating adult central nervous system.

Adult mammalian optic nerve axons are able to regenerate, when provided with the permissive environment of an autologous peripheral nerve graft, which is usually the sciatic nerve. This study demonstrates the ability of adult rat optic nerve axons to regenerate through the preformed perforations of a polyimide electrode carrier implanted at the interface between the proximal stump of the cut optic nerve and the stump of the peripheral nerve piece used for grafting. Evidence that retinal ganglion cells regenerated their axons through the perforated electrode carrier was obtained by retrograde labeling with a fluorescent dye deposited into the sciatic nerve graft beyond the nerve-carrier-nerve junction. The number of regenerating cells could be enhanced by injecting neuroprotective drugs like aurintricarboxylic acid and cortisol intravitreally. A second line of evidence was obtained by immunohistochemical staining with antibodies to neurofilament. Third, electrical activity of the regenerating nerves was recorded after stimulating the retina with a flash of light. The results suggest that a regenerating central nerve tract may serve as an experimental model to implant artificial microdevices to monitor the physiological and topographical properties of neurites passing through the device or to stimulate them, thus interfering with their potential to grow. This study reports for the first time that the optic nerve has unique properties, which aids in the realization of these goals.

Animals↗

PubMed2003

Radiation-induced modulation of the microglial population in the normal and injured mature spinal cord.

Recent attempts by other investigators to enhance repair processes in the spinal cord have involved the administration of X rays to spinal cord injury sites. Although some functional improvement has been reported, the underlying cellular changes within the irradiated spinal cords are not clear. Studies initiated recently in this laboratory examined the potential of X rays to modulate nonneuronal cell populations associated with an injury site in adult mammalian spinal cords. These studies revealed a unique and previously unreported radiosensitivity of the microglial cell population. Administration of X radiation to a unilateral dorsal lesion cavity in the cervical spinal cord revealed a significant decrease (approximately half) in numbers of microglia associated with the cavity. Even more unexpected were the significant decreases in microglial cells observed on the nonlesioned side of the spinal cord or in sham-operated spinal cords in irradiated rats. In contrast to reports of others, densitometric quantification of GFAP immunoreactive cells and processes indicated no differences in the astrocytic reactions associated with the lesion cavities between nonirradiated and irradiated groups in our studies. The demonstration that exposure of a spinal cord injury site to radiation modifies the responses of certain components of the glial environment to injury may offer a noninvasive approach for direct treatment of that site. Studies are in progress to determine if this altered glial environment enhances the extension of regrowing axons from a peripheral nerve graft across the interface with the irradiated lesion cavity and into the spinal cord parenchyma.

Age Factors↗

PubMedDate not supplied

Characterization of Mannheimia (Pasteurella) haemolytica leukotoxin interaction with bovine alveolar macrophage beta2 integrins.

Mannheimia (Pasteurella) haemolytica, the etiologic agent of bovine pneumonic mannheimiosis, produces an exotoxic leukotoxin. The leukotoxin (LktA) is a member of the RTX (repeats in toxin) family of bacterial cytolysins and is distinguished from other toxins by its unique target cell specificity to ruminant leukocytes occurring through binding to a specific receptor. We have demonstrated previously that the beta2 integrin LFA-1 is a receptor for LktA in bovine leukocytes and is involved in leukotoxin-induced biological effects. However the subunits within LFA-1 involved in binding to LktA, and post-binding signaling leading to cellular activation have not been well characterized. The purpose of our study was to pinpoint these precise subunits on bovine alveolar macrophages and to characterize their interaction with LktA. The results in this study indicate that although LktA can efficiently bind to the CD18 subunit of both LFA-1 and Mac-1, post-binding signaling events including elevation of intracellular calcium and CD18 tail phosphorylation are only observed through LFA-1. Furthermore, LktA also binds to the CD11a subunit of LFA-1. LktA binding to CD11a could be inhibited by a small molecule inhibitor of the I(inserted)-domain, the major ligand binding interface on CD11a. I-domain inhibition significantly blunts LktA-induced intracellular calcium elevation and tyrosine phosphorylation of the CD18 tail. Based on our results we suggest that LFA-1 serves as the functional leukotoxin receptor on bovine alveolar macrophages.

Animals↗

PubMed2005

Asymmetric interaction between rod cyclic GMP phosphodiesterase gamma subunits and alphabeta subunits.

Rod phosphodiesterase (PDE6) is the central effector enzyme in vertebrate visual transduction. Holo-PDE6 consists of two similar catalytic subunits (Palphabeta) and two identical inhibitory subunits (Pgamma). Palphabeta is the only heterodimer in the PDE superfamily, yet its significance for the function of PDE6 is poorly understood. An unequal interaction of Pgamma with Pbeta as compared with Palpha in the PDE6 complex has not been reported. We investigated the interaction interface between full-length Pgamma and Palphabeta, by differentiating Pgamma interaction with each individual Palphabeta subunit through radiolabel transfer from various positions throughout the entire Pgamma molecule. The efficiency of radiolabel transfer indicates that the close vicinity of serine 40 on Pgamma makes a major contribution to the interaction with Palphabeta. In addition, a striking asymmetry of interaction between the Pgamma polycationic region and the Palphabeta subunits was observed when the stoichiometry of Pgamma versus the Palphabeta dimer was below 2. Preferential photolabeling on Pbeta from Pgamma position 40 and on Palpha from position 30 increased while lowering the Pgamma/Palphabeta ratio, but diminished when the Pgamma/Palphabeta ratio was over 2. Our finding leads to the conclusion that two classes of Pgamma binding sites exist on Palphabeta, each composed of GAF domains in both Palpha and Pbeta, differing from the conventional models suggesting that each Pgamma binds only one of the Palphabeta catalytic subunits. This new model leads to insight into how the unique Palphabeta heterodimer contributes to the sophisticated regulation in visual transduction through interaction with Pgamma.

3',5'-Cyclic-GMP Phosphodiesterases↗

PubMed2004

CleanEx: a database of heterogeneous gene expression data based on a consistent gene nomenclature.

The main goal of CleanEx is to provide access to public gene expression data via unique gene names. A second objective is to represent heterogeneous expression data produced by different technologies in a way that facilitates joint analysis and cross-data set comparisons. A consistent and up-to-date gene nomenclature is achieved by associating each single experiment with a permanent target identifier consisting of a physical description of the targeted RNA population or the hybridization reagent used. These targets are then mapped at regular intervals to the growing and evolving catalogues of human genes and genes from model organisms. The completely automatic mapping procedure relies partly on external genome information resources such as UniGene and RefSeq. The central part of CleanEx is a weekly built gene index containing cross-references to all public expression data already incorporated into the system. In addition, the expression target database of CleanEx provides gene mapping and quality control information for various types of experimental resource, such as cDNA clones or Affymetrix probe sets. The web-based query interfaces offer access to individual entries via text string searches or quantitative expression criteria. CleanEx is accessible at: http://www.cleanex.isb-sib.ch/.

Animals↗

PubMed1999

The Mouse Genome Database (MGD): genetic and genomic information about the laboratory mouse. The Mouse Genome Database Group.

The Mouse Genome Database (MGD) focuses on the integration of mapping, homology, polymorphism and molecular data about the laboratory mouse. Detailed descriptions of genes including their chromosomal location, gene function, disease associations, mutant phenotypes, molecular polymorphisms and links to representative sequences including ESTs are integrated within MGD. The association of information from experiment to gene to genome requires careful coordination and implementation of standardized vocabularies, unique nomenclature constructions, and detailed information derived from multiple sources. This information is linked to other public databases that focus on additional information such as expression patterns, sequences, bibliographic details and large mapping panel data. Scientists participate in the curation of MGD data by generating the Chromosome Committee Reports, consulting on gene family nomenclature revisions, and providing descriptions of mouse strain characteristics and of new mutant phenotypes. MGD is accessible at http://www.informatics.jax.org

Animals↗

PubMed1998

Recovery of function processes in human amnesia: evidence from transient global amnesia.

There are few clues as to the processes that underlie recovery of function from human amnesia. Evidence is offered from the perspective of a study of recovery of function during an episode of transient global amnesia (TGA) that occurred as a complication of a cerebral angiographic procedure being carried out in a neurosciences centre, and where there was therefore a unique opportunity to examine acute changes in memory function. This allowed us to conduct the first quantitative study where shrinkage of anterograde and retrograde memory loss was plotted at four separate intervals throughout the acute recovery process, and also 24 hr later. Recovery of retrograde amnesia preceded recovery from anterograde amnesia. Resolution of a naming deficit more closely paralleled recovery from retrograde amnesia rather than anterograde amnesia. Within retrograde amnesia for public events, there was a temporal gradient of memory loss, with more recent events affected to a greater degree than earlier events. Within anterograde amnesia, picture recognition memory preceded recovery of story recall memory. On the basis of these findings, and related observations in the published literature, it is proposed that recovery from some types of human amnesia, such as that associated with TGA, follows a 'lateral-to-medial' rule--lateral inferotemporal areas that play a major role in retrograde amnesia recover first from hypometabolism related to the TGA attack, followed by 'interface' areas such as the rhinal and parahippocampal cortices that are considered to have a role in both anterograde and retrograde memory functioning, with the last areas to recover physiological integrity being discrete limbic-diencephalic structures such as the hippocampus.

Adult↗

PubMed1999

Substrate-adsorbed collagen and cell secreted fibronectin concertedly induce cell migration on poly(lactide-glycolide) substrates.

Limited epithelial cell migration on synthetic polymeric biomaterials, such as polyesters, presents a serious challenge to their use as scaffolds for artificial skin analogs. The mechanisms by which a physiologic matrix interface on such polymers may regulate and promote cell migration under 'activated conditions' were the focus of this study. We have quantified the migration behavior of epidermal growth factor (EGF) stimulated epidermal keratinocytes on 50:50 poly-D,L(lactide-glycolide) (PLGA) substrates, following exogenous and cell-derived substrate conditioning based on the model matrix proteins, collagen and fibronectin. We report that 'non-conditioned' PLGA substrates elicited poor levels of keratinocyte migration. However, keratinocyte migration was significantly enhanced upon the adsorption of type I collagen, and was only weakly enhanced with fibronectin adsorption. Molecular analysis of the mechanism of enhanced migration on collagen-PLGA substrates showed that keratinocyte migration was sensitive to cell-derived fibronectin conditioning, but not to cell-secreted collagen conditioning. Fibronectin control of cell migration on collagen-PLGA was found to be both stoichiometric and biologically specific, mediated via adhesion involving keratinocyte alpha v integrin receptors. Based on our results, we propose a unique paradigm for induction of cell migration on a non-physiologic synthetic polymer using concerted interactions between primary, polymer-instructed matrix remodeling and secondary, cell-derived matrix remodeling.

Adsorption↗

PubMed2006

Microfluidic liquid chromatography system for proteomic applications and biomarker screening.

A microfluidic liquid chromatography (LC) system for proteomic investigations that integrates all the necessary components for stand-alone operation, i.e., pump, valve, separation column, and electrospray interface, is described in this paper. The overall size of the LC device is small enough to enable the integration of two fully functional separation systems on a 3 in. x 1 in. glass microchip. A multichannel architecture that uses electroosmotic pumping principles provides the necessary functionality for eluent propulsion and sample valving. The flow rates generated within these chips are fully consistent with the requirements of nano-LC platforms that are routinely used in proteomic applications. The microfluidic device was evaluated for the analysis of a protein digest obtained from the MCF7 breast cancer cell line. The cytosolic protein extract was processed according to a shotgun protocol, and after tryptic digestion and prefractionation using strong cation exchange chromatography (SCX), selected sample subfractions were analyzed with conventional and microfluidic LC platforms. Using similar experimental conditions, the performance of the microchip LC was comparable to that obtained with benchtop instrumentation, providing an overlap of 75% in proteins that were identified by more than two unique peptides. The microfluidic LC analysis of a protein-rich SCX fraction enabled the confident identification of 77 proteins by using conventional data filtering parameters, of 39 proteins with p < 0.001, and of 5 proteins that are known to be cancer-specific biomarkers, demonstrating thus the potential applicability of these chips for future high-throughput biomarker screening applications.

Biomarkers, Tumor↗

PubMed2000

Identification of monoclonal nonspecific suppressor factor beta (mNSFbeta) as one of the genes differentially expressed at implantation sites compared to interimplantation sites in the mouse uterus.

Successful implantation requires synchronous development of and active dialogue between the maternal endometrium and the implanting blastocyst. While it is well established that appropriate maternal steroid hormones are essential for endometrial preparation for implantation, the molecular events at the actual site of implantation are still little understood. The aims of our studies were to identify genes explicitly expressed or repressed at the sites of implantation by utilising RNA differential display (DDPCR), and to establish the roles of these genes in the implantation process in a mouse model. Ten bands unique in implantation sites compared to interimplantation sites were identified by DDPCR and subsequently confirmed by Northern blotting. One of these bands contained a cDNA fragment that was highly homologous to mouse monoclonal nonspecific suppressor factor beta (MNSFbeta) or Fau. The full cDNA sequence of this gene, obtained by screening a lambdagt11 cDNA library, was essentially the same as MNSFbeta, except that it had much longer 5' untranslated region. Interestingly, both Northern and immunohistochemical analysis showed that the expression of this gene was much lower in implantation sites compared to interimplantation sites on day 4.5 of pregnancy, when embryos first attach to the uterus and initiate implantation, and on day 5.5, when implantation has advanced. These results suggest a role for MNSF during implantation and early pregnancy, possibly through regulating the proliferation and/or differentiation of uterine stromal cells. It may also be involved in the selective production of TH2-type cytokines in implantation sites to regulate the immune system at the maternal-fetal interface.

Amino Acid Sequence↗

PubMed2005

The blood-testis barrier: its biology, regulation, and physiological role in spermatogenesis.

The blood-testis barrier (BTB) in mammals, such as rats, is composed of the tight junction (TJ), the basal ectoplasmic specialization (basal ES), the basal tubulobulbar complex (basal TBC) (both are testis-specific actin-based adherens junction [AJ] types), and the desmosome-like junction that are present side-by-side in the seminiferous epithelium. The BTB physically divides the seminiferous epithelium into basal and apical (or adluminal) compartments, and is pivotal to spermatogenesis. Besides its function as an immunological barrier to segregate the postmeiotic germ-cell antigens from the systemic circulation, it creates a unique microenvironment for germ-cell development and confers cell polarity. During spermatogenesis, the BTB in rodents must physically disassemble to permit the passage of preleptotene and leptotene spermatocytes. This occurs at late stage VII through early stage VIII of the epithelial cycle. Studies have shown that this dynamic BTB restructuring to facilitate germ-cell migration is regulated by two cytokines, namely transforming growth factor-beta3 (TGF-beta3) and tumor necrosis factor-alpha (TNFalpha), via downstream mitogen-activated protein kinases. These cytokines determine the homeostasis of TJ- and basal ES-structural proteins, proteases, protease inhibitors, and other extracellular matrix (ECM) proteins (e.g., collagen) in the seminiferous epithelium. Some of these molecules are known regulators of focal contacts between the ECM and other actively migrating cells, such as macrophages, fibroblasts, or malignant cells. These findings also illustrate that cell-cell junction restructuring at the BTB is regulated by mechanisms involved in the junction turnover at the cell-matrix interface. This review critically discusses these latest findings in the field in light of their significance in the biology and regulation of the BTB pertinent to spermatogenesis.

Animals↗

PubMed1998

Active monomeric and dimeric forms of Pseudomonas putida glyoxalase I: evidence for 3D domain swapping.

3D domain swapping of proteins involves the interconversion of a monomer containing a single domain-domain interface and a 2-fold symmetrical dimer containing two equivalent intermolecular interfaces. Human glyoxalase I has the structure of a domain-swapped dimer [Cameron, A. D., Olin, B., Ridderström, M., Mannervik, B., and Jones, T. A. (1997) EMBO J. 16, 3386-3395] but Pseudomonas putida glyoxalase I has been reported to be monomeric [Rhee, H.-I., Murata, K., and Kimura, A. (1986) Biochem. Biophys. Res. Commun. 141, 993-999]. We show here that recombinant P. putida glyoxalase I is an active dimer (kcat approximately 500 +/- 100 s-1; KM approximately 0.4 +/- 0.2 mM) with two zinc ions per dimer. The zinc is required for structure and function. However, treatment of the dimer with glutathione yields an active monomer (kcat approximately 115 +/- 40 s-1; KM approximately 1.4 +/- 0.4 mM) containing a single zinc ion. The monomer is metastable and slowly reverts to the active dimer in the absence of glutathione. Thus, glyoxalase I appears to be a novel example of a single protein able to exist in two alternative domain-swapped forms. It is unique among domain-swapped proteins in that the active site and an essential metal binding site are apparently disassembled and reassembled by the process of domain swapping. Furthermore, it is the only example to date in which 3D domain swapping can be regulated by a small organic ligand.

Apoenzymes↗

PubMed2004

Microemulsification of triglyceride sebum and the role of interfacial structure on bicontinuous phase behavior.

A unique triblock surfactant is reported that allows for the efficient microemulsification of triglycerides. Unlike the results of all previous efforts, these triglyceride microemulsions can be formed without the use of cosurfactants or dilution with co-oils and follow the classical patterns of surfactant phase behavior exhibited by mixtures of water, alkane oils, and nonionic oligoethylene glycol surfactants, i.e., progression from oil/water emulsions to one-phase microemulsions to water/oil emulsions with increasing temperature. Lamellar phases that usually dominate the aqueous phase behavior of surfactant/triglyceride mixtures are suppressed, allowing for the formation of single-phase microemulsions containing equal amounts of triglyceride and water. These isotropic and low-viscous fluids are particularly useful for cleansing and delivery of functional ingredients in skin care products. The effects of mixing a variety of typical skin care ingredients and components of sebum (skin oil) were also explored. Fatty acids significantly reduce the average microemulsion temperature, while other ingredients and oils, which do not partition at the oil/water interface, have less impact on the phase behavior. In all cases, one-phase microemulsions containing equal amounts of oil and water can be formed even at high additive concentrations. Indeed, partial replacement oftriglyceride with any of the additives examined consistently reduced the amount of surfactant necessary to form single-phase microemulsions. However, the greatest boost in surfactant efficiency was found with the addition of medium molecular weight amphiphilic block copolymers.

Emulsions↗

PubMedDate not supplied

Using Internet discussion boards as virtual focus groups.

While the potential of Internet-based qualitative research methods is substantial, such methods are not without their problems. Some of these methodological challenges are unique to the medium, while others are similar to those of more traditional qualitative methods. This article presents some of these methodological challenges, and explores some of the issues involved in using on-line discussion boards as virtual focus groups in a study of perimenopausal women with migraines. Design of the study and its advantages and disadvantages are discussed, including the role of the moderator. Some of the problems encountered included potential for misunderstandings due to limits of written communications, and difficulty encouraging participation.

Climacteric↗

PubMed2002

Unique assignment of inter-subunit association in GABA(A) alpha 1 beta 3 gamma 2 receptors determined by molecular modeling.

Recent publications defined requirements for inter-subunit contacts in a benzodiazepine-sensitive GABA(A) receptor (GABA(A)R alpha 1 beta 3 gamma 2). There is strong evidence that the heteropentameric receptor contains two alpha 1, two beta 3, and one gamma 2 subunit. However, the available data do not distinguish two possibilities: When viewed clockwise from an extracellular viewpoint the subunits could be arranged in either gamma 2 beta 3 alpha 1 beta 3 alpha 1 or gamma 2 alpha 1 beta 3 alpha 1 beta 3 configurations. Here we use molecular modeling to thread the relevant GABA(A)R subunit sequences onto a template of homopentameric subunits in the crystal structure of the acetylcholine binding protein (AChBP). The GABA(A) sequences are known to have 15-18% identity with the acetylcholine binding protein and nearly all residues that are conserved within the nAChR family are present in AChBP. The correctly aligned GABA(A) sequences were threaded onto the AChBP template in the gamma 2 beta 3 alpha 1 beta 3 alpha 1 or gamma 2 alpha 1 beta 3 alpha 1 beta 3 arrangements. Only the gamma 2 alpha 1 beta 3 alpha 1 beta 3 arrangement satisfied three known criteria: (1) alpha 1 His(102) binds at the gamma 2 subunit interface in proximity to gamma 2 residues Thr(142), Phe(77), and Met(130); (2) alpha 1 residues 80-100 bind near gamma 2 residues 91-104; and (3) alpha 1 residues 58-67 bind near the beta 3 subunit interface. In addition to predicting the most likely inter-subunit arrangement, the model predicts which residues form the GABA and benzodiazepine binding sites.

Amino Acid Sequence↗

PubMed2005

The folding landscape of an alpha-lytic protease variant reveals the role of a conserved beta-hairpin in the development of kinetic stability.

Most secreted bacterial proteases, including alpha-lytic protease (alphaLP), are synthesized with covalently attached pro regions necessary for their folding. The alphaLP folding landscape revealed that its pro region, a potent folding catalyst, is required to circumvent an extremely large folding free energy of activation that appears to be a consequence of its unique unfolding transition. Remarkably, the alphaLP native state is thermodynamically unstable; a large unfolding free energy barrier is solely responsible for the persistence of its native state. Although alphaLP folding is well characterized, the structural origins of its remarkable folding mechanism remain unclear. A conserved beta-hairpin in the C-terminal domain was identified as a structural element whose formation and positioning may contribute to the large folding free energy barrier. In this article, we characterize the folding of an alphaLP variant with a more favorable beta-hairpin turn conformation (alphaLP(beta-turn)). Indeed, alphaLP(beta-turn) pro region-catalyzed folding is faster than that for alphaLP. However, instead of accelerating spontaneous folding, alphaLP(beta-turn) actually unfolds more slowly than alphaLP. Our data support a model where the beta-hairpin is formed early, but its packing with a loop in the N-terminal domain happens late in the folding reaction. This tight packing at the domain interface enhances the kinetic stability of alphaLP(beta-turn), to nearly the same degree as the change between alphaLP and a faster folding homolog. However, alphaLP(beta-turn) has impaired proteolytic activity that negates the beneficial folding properties of this variant. This study demonstrates the evolutionary limitations imposed by the simultaneous optimization of folding and functional properties.

Bacterial Proteins↗

PubMed2001

Vitamin C enhances differentiation of a continuous keratinocyte cell line (REK) into epidermis with normal stratum corneum ultrastructure and functional permeability barrier.

A continuous rat epidermal cell line (rat epidermal keratinocyte; REK) formed a morphologically well-organized epidermis in the absence of feeder cells when grown for 3 weeks on a collagen gel in culture inserts at an air-liquid interface, and developed a permeability barrier resembling that of human skin. By 2 weeks, an orthokeratinized epidermis evolved with the suprabasal layers exhibiting the differentiation markers keratin 10, involucrin, and filaggrin. Granular cells with keratohyalin granules and lamellar bodies, and corneocytes with cornified envelopes and tightly packed keratin filaments were present. Morphologically, vitamin C supplementation of the culture further enhanced the normal wavy pattern of the stratum corneum, the number of keratohyalin granules present, and the quantity and organization of intercellular lipid lamellae in the interstices of the stratum corneum. The morphological enhancements observed with vitamin C correlated with improved epidermal barrier function, as indicated by reduction of the permeation rates of tritiated corticosterone and mannitol, and transepidermal water loss, with values close to those of human skin. Moreover, filaggrin mRNA was increased by vitamin C, and western blots confirmed higher levels of profilaggrin and filaggrin, suggesting that vitamin C also influences keratinocyte differentiation in aspects other than the synthesis and organization of barrier lipids. The unique REK cell line in organotypic culture thus provides an easily maintained and reproducible model for studies on epidermal differentiation and transepidermal permeation.

Animals↗

PubMed2004

Model system for growing and quantifying Streptococcus pneumoniae biofilms in situ and in real time.

Streptococcus pneumoniae forms biofilms, but little is known about its extracellular polymeric substances (EPS) or the kinetics of biofilm formation. A system was developed to enable the simultaneous measurement of cells and the EPS of biofilm-associated S. pneumoniae in situ over time. A biofilm reactor containing germanium coupons was interfaced to an attenuated total reflectance (ATR) germanium cell of a Fourier transform infrared (FTIR) laser spectrometer. Biofilm-associated cells were recovered from the coupons and quantified by total and viable cell count methods. ATR-FTIR spectroscopy of biofilms formed on the germanium internal reflection element (IRE) of the ATR cell provided a continuous spectrum of biofilm protein and polysaccharide (a measure of the EPS). Staining of the biofilms on the IRE surface with specific fluorescent probes provided confirmatory evidence for the biofilm structure and the presence of biofilm polysaccharides. Biofilm protein and polysaccharides were detected within hours after inoculation and continued to increase for the next 141 h. The polysaccharide band increased at a substantially higher rate than did the protein band, demonstrating increasing coverage of the IRE surface with biofilm polysaccharides. The biofilm total cell counts on germanium coupons stabilized after 21 h, at approximately 10(5) cells per cm(2), while viable counts decreased as the biofilm aged. This system is unique in its ability to detect and quantify biofilm-associated cells and EPS of S. pneumoniae over time by using multiple, corroborative techniques. This approach could prove useful for the study of biofilm processes of this or other microorganisms of clinical or industrial relevance.

Bacteriological Techniques↗