Development of a Rapid Streptavidin Capture-Based Assay for the Tyrosine Phosphorylated CSF-1R in Peripheral Blood Mononuclear Cells

A novel assay was developed to measure ratio of p-FMS (phospho FMS) to FMS using the Meso Scale Discovery^® (MSD) technology and compared to the routinely used, IP-Western based approach. The existing IP-Western assay used lysed PBMCs (Peripheral Blood Mononuclear Cells) that were immunoprecipitated (IP) overnight, and assayed qualitatively by Western analysis. This procedure takes three days for completion. The novel IP-MSD method described in this paper employed immunoprecipitation of the samples for one hour, followed by assessment of the samples by a ruthenium labeled secondary antibody on a 96-well Streptavidin-coated MSD plate. This IP-MSD method was semi-quantitative, could be run in less than a day, required one-eighth the volume of sample, and compared well to the IP-Western method. In order to measure p-FMS/FMS, samples from healthy volunteers (HV) were first stimulated with CSF-1(Macrophage colony-stimulating factor) to initiate the changes in the phosphotyrosyl signaling complexes in FMS. The objective of the present work was to develop a high throughput assay that measured p-FMS/FMS semi-quantitatively, with minimal sample requirement, and most importantly compared well to the current IP-Western assay.     

A critical physiological role of zinc in the structure and function of biomembranes

Of the large number of recognized zinc metalloenzymes, the activities of only a few are significantly decreased in severely zinc deficient animals. On the other hand, physiological pathology is manifested rapidly after dietary zinc deprivation. This shows that zinc exerts physiological and biochemical roles other than as a component of the known zinc metalloenzymes. The research results reviewed here suggest that zinc plays an important role in the maintenance of membrane structure and function.     

Structural definition of the non-reducing termini of mannose-capped LAM from Mycobacterium tuberculosis through selective enzymatic degradation and fast atom bombardment-mass spectrometry

The application of extracellular arabinases from aCellulomonassp. and fast atom bombardment-mass spectrometry (FAB-MS) providednew insight into the structure of lipoarabinomannan (LAM) ofMycobacterium tuberculosis, a key molecule in the pathogenesisand physiology of the tubercle bacillus. Previously, the non-reducingarabinan ends of LAM from the virulent (Erdman) strain of M.tuberculosiswere shown to be ‘capped’ by short (a1     

Cellular pathophysiology of cystic kidney disease: insight into future therapies.

Polycystic kidney disease (PKD) is a developmental kidney disorder which can be inherited as either an autosomal dominant trait, with an incidence of 1:50 to 1:1000, or as an autosomal recessive trait with an incidence of 1:6,000 to 1:40,000. Three different genes have now been cloned that are associated with mutations that cause PKD. Two of these are linked to the most common forms of the dominant disease while the third is associated with the orpk mouse model of recessive polycystic kidney disease. Advances in understanding the molecular genetics of PKD have been paralleled by new insights into the cellular pathophysiology of cyst formation and progressive enlargement. Current data suggest that a number of PKD proteins may interact in a complex, which when disrupted by mutations in PKD genes may lead to altered epithelial proliferative activity, secretion, and cell matrix biology. The identification of a unique cystic epithelial phenotype presents new opportunities for targeted therapies. These include targeted gene therapy, gene complementation, and specific immunological or pharmacological interruption of growth factor pathways.     

Experimental investigation of the scaphoid strain during wrist motion

The scaphoid is the most frequently fractured of the carpal bones [Taleisnik, J., The Wrist, Churchill Livingstone, New York (1985)]. This project was undertaken to qualitatively evaluate the strain in the scaphoid during wrist motion using a newly developed strain gage method. Strain gage resettes were mounted within the scaphoid and the range of motion of the hand was monitored using a custom designed electrogoniometer and data acquisition system. Ten specimens were utilized for this study. Results indicated that supination/pronation (S/P) of the forearm did not affect the strain in the scaphoid. A map of the strain in the waist of the scaphoid, as a function of flexion/extension (F/E) and radial/ulnar deviation (R/U), was generated. The contour plot of scaphoid strain vs range of motion (ROM) shows a valley where strains are low. Minimum scaphoid strain was found near neutral F/E and 15° of ulnar deviation.