Suction blister fluid as potential body fluid for biomarker proteins

Early diagnosis is important for effective disease management. Measurement of biomarkers present at the local level of the skin could be advantageous in facilitating the diagnostic process. The analysis of the proteome of suction blister fluid, representative for the interstitial fluid of the skin, is therefore a desirable first step in the search for potential biomarkers involved in biological pathways of particular diseases. Here, we describe a global analysis of the suction blister fluid proteome as potential body fluid for biomarker proteins. The suction blister fluid proteome was compared with a serum proteome analyzed using identical protocols. By using stringent criteria allowing less than 1% false positive identifications, we were able to detect, using identical experimental conditions and amount of starting material, 401 proteins in suction blister fluid and 240 proteins in serum. As a major result of our analysis we construct a prejudiced list of 34 proteins, relatively highly and uniquely detected in suction blister fluid as compared to serum, with established and putative characteristics as biomarkers. We conclude that suction blister fluid might potentially serve as a good alternative biomarker body fluid for diseases that involve the skin.     

Interstitial fluid lipoproteins

While a wide variety of techniques has been used to collect samples of interstitial fluid, most of our detailed knowledge about the composition of interstitial fluid lipoproteins has come from lymph collection studies. The considerable variability of lymph data probably reflects the effect of variable metabolic modification and different capillary permeabilities on the lipoprotein composition of interstitial fluid. All density classes of plasma lipoproteins are present in lymph. In peripheral lymph, the lymph/plasma concentration ratios of lipoproteins vary from 0.03 for VLDL-sized particles to 0.2 for HDL. Lymph from more permeable vascular beds, such as lung and myocardium, contains proportionately more lipoproteins. Their lymph/plasma concentration ratios vary from 0.1 to 0.6. In general, lymph lipoproteins are more heterogeneous in size than their plasma counterparts. Lymph HDL and LDL contain larger and smaller particles than their plasma equivalents. Lymph lipoproteins have unusual shapes (square packing and discoidal), chemical compositions, and molecular charge, which suggest de novo formation and/or extensive peripheral modification. Lymph HDL and LDL are enriched in free cholesterol. Lymph HDL also has increased cholesterol/protein and phospholipid/protein (especially sphingomyelin) ratios (Sloop, C.H., L. Dory, and P.S. Roheim, unpublished observations). Lymph HDL apoprotein composition differs from that of plasma, with an increase in apoE and apoA-IV content relative to apoA-I. These discoidal HDL particles may be products of an initial stage of reverse cholesterol transport. We believe further study of their metabolic fate would give important information concerning the later stages of reverse cholesterol transport.     

A bioseparation apparatus with high-pressure fluid injection and fluid sampling

A novel apparatus in which fluids may be injected and sampled at high pressure is described. Bioseparation applications of the apparatus were demonstrated in three model systems: (1) lambdaDNA was eluted under pressure from an anion exchange column into a low-salt (0.25 M) buffer, thereby eliminating conventional time-consuming desalting procedures required for downstream analysis of the DNA; (2) RNA was separated under pressure from a RNA/DNA mixture, thereby enabling rapid differential preparation of nucleic acids; and (3) an antibody was purified from a protein mixture by affinity capture at one pressure and dissociation from the antigen binding partner at a second pressure, thereby enabling the immunoreactivities of both antibody and antigen to be preserved during the separation process.     

Bounding fluid viscosity and translational diffusion in a fluid lipid bilayer

Fluorescence recovery after photobleaching was used to investigate the translational diffusion of a fluorescent derivative of a membrane-spanning lipid in L phase multibilayers of 1-palmitoyl-2-oleoylphosphatidylcholine prepared in water and in glycerol. The translational diffusion coefficient in hydrated bilayers (D _w) ranged between 2 and 5x10^–8 cm²/s and in glycerinated bilayers (D _g) the range was between 3 and 24×10^–10 cm²/s between 10° and 40°C. These results are discussed in terms of models for diffusion in membranes.     

Creatine kinase and ATPase in human seminal fluid and prostatic fluid

A creatine kinase assay based on estimation of creatine liberated from creatine phosphate was accurate and reproducible for use with seminal or prostatic fluid, after allowance was made for acid phosphatase interference. Comparison of this method with one which relies on enzymic coupling of ATP formation to NADP+ oxidation shows that the latter under-estimates creatine kinase activity by a factor of about 3. This discrepancy could be due to the high ATPase activity found in prostatic and seminal fluid. Uncritical use of the NADP+ assay might account for different seminal creatine kinase values reported in the literature. Interrelationships between ATPase, creatine kinase and zinc suggest that seminal ATPase is a prostatic secretory product while creatine kinase may be multiglandular in origin.     

Comparative study of the fluid dynamics of bottom spray fluid bed coaters

Fluid dynamics of pellets processed in bottom spray traditional Wurster coating and swirl accelerated air (precision) coating were compared with the intent to understand and facilitate improvements in the coating processes. Fluid dynamics was described by pellet mass flow rate (MFR) obtained using a pellet collection system and images captured using high speed photography. Pellet flow within the partition column was found to be denser and slower in Wurster coating than in precision coating, suggesting a higher tendency of agglomeration during the coating process. The influence of partition gap and load on the MFR indicated that the mechanism of transport of pellets into the coating zone in precision coating depended on a strong suction, whereas in Wurster coating, pellets were transported by a combination of peripheral fluidization, gravity, and weak suction pressure. In precision coating, MFR was found to increase uniformly with air flow rate and atomizing pressure, whereas MFR in Wurster coating did not correlate as well with air flow rate and atomizing pressure. This demonstration showed that transport in precision coating was air dominated. In conclusion, fluid dynamics in precision coating was found to be air dominated and dependent on pressure differential, thus it is more responsive to changes in operational variables than Wurster coating.     

Rheology of synovial fluid

After a discussion of the role of synovial fluid as a joint lubricant, rheological measurements are described with both normal (healthy) synovial fluids and pathological ones. Shear stress and first normal stress difference are measured as a function of shear gradient to calculate the apparent shear viscosity eta 1 and the apparent normal viscosity psi 7 as well as an apparent shear modulus G'. It is found, that in case of diseased synoviae all rheological parameters deteriorate. Most significant changes are observed with the zero shear viscosity eta 0, the shear modulus G', and a characteristic time theta 1, which is the reciprocal of the critical shear rate Dc which determines the onset of shear thinning. The rheological deterioration of synovial fluids is explained in terms of solute structure, whereby a molecular mass of the backbone hyaluronic acid of at least 10(7) g.mol-1 is required for satisfactory function. A theory of the rheological performance of normal synovial fluid as well as its pathological deterioration is proposed.     

The excitable fluid mosaic

The Fluid Mosaic Model by Singer & Nicolson proposes that biological membranes consist of a fluid lipid layer into which integral proteins are embedded. The lipid membrane acts as a two-dimensional liquid in which the proteins can diffuse and interact. Until today, this view seems very reasonable and is the predominant picture in the literature. However, there exist broad melting transitions in biomembranes some 10–20 degrees below physiological temperatures that reach up to body temperature. Since they are found below body temperature, Singer & Nicolson did not pay any further attention to the melting process. But this is a valid view only as long as nothing happens. The transition temperature can be influenced by membrane tension, pH, ionic strength and other variables. Therefore, it is not generally correct that the physiological temperature is above this transition. The control over the membrane state by changing the intensive variables renders the membrane as a whole excitable. One expects phase behavior and domain formation that leads to protein sorting and changes in membrane function. Thus, the lipids become an active ingredient of the biological membrane. The melting transition affects the elastic constants of the membrane. This allows for the generation of propagating pulses in nerves and the formation of ion-channel-like pores in the lipid membranes. Here we show that on top of the fluid mosaic concept there exists a wealth of excitable phenomena that go beyond the original picture of Singer & Nicolson.¹     

Automated cerebrospinal fluid cytology

OBJECTIVE: To evaluate the Abbott CELL-DYN Sapphire cytometer for cerebrospinal fluid (CSF) cell count and differentiation. METHODS: One hundred three analyses of CSF cells by the CELL-DYN Sapphire were compared with routine cell count and microscopic differentiation and correlation coefficients calculated. RESULTS: The total cell count of both methods correlated well. The detection of erythrocytes was good (0.898), and a higher content of erythrocytes >100/microL had little effect on total leukocyte count. The correlation between both methods was best with higher leukocyte counts >25/microL (r=0.987), whereas at cell counts <25/microL, the correlation was considerably less precise (r=0.613). For the differentiation of cells, lymphocytes and neutrophils showed moderate correlation. The results for monocytes and eosinophils did not correlate. CONCLUSION: The results for the total cell count in this study are comparable with those achieved with the Bayer Advia 120. While the Abbott CELL-DYN Sapphire yielded slightly better results for erythrocytes and total cell count with a higher erythrocyte content, the Advia 120 achieved slightly better results of lymphocyte and neutrophil count in a previous study.