Laser Raman investigation of the conformation of human immunoglobulin G.

Laser Raman spectra of human immunoglobulin G in neutral solution, as well as in the lyophilized and alkaline-denatured states are presented. In the spectrum of the native protein, the amide III band appears at 1240 cm-1 and is assigned to the presence of beta-sheet structure. From its intensity, using a procedure described in this paper, we evaluate the beta-structure content to 37 +/- 4%. This result is supported by the strong amide I' band at 1667 cm-1 and by the presence in the spectra of two bands at 991 and 1078 cm-1, respectively assigned to the C-C and C-N skeletal stretching modes. The differences between the spectrum of the lyophilized powder and that of the solution show that the lyophilization process induces conformational changes that perturb the local environment of some of the tryptophan residues and alter the secondary structure of immunoglobulin G. The beta-structure appears to be more uniform and more abundant in solution. When the protein is denatured at pH 11, the amide III and amide I'bands, which become weaker and broader, shift in frequency from 1240 to 1248 cm-1 and from 1667 to 1656 cm-1 respectively. These changes indicate a decrease in the amount of beta-structure and a transition toward a much more disordered conformation. During the denaturation, the intensities of many bands of the aromatic chromophores change, notably the tryptophan peaks at 879, 1359 and 1573 cm-1.

Evaluation of a simplified filtration technique for the routine measurement of erythrocyte deformability.

The purpose of this study was to assess the value of a widely-used simplified filtration technique (mini-test) by comparing it, in parallel experiments, with a constant-flow rate method currently used in our laboratory for the measurement of erythrocyte deformability. Both methods employed 3 micrometer pore-diameter Nucleopore membranes and were performed under different flow conditions, using washed erythrocyte suspensions. In a total of 49 evaluable test pairs obtained from blood samples of 55 patients with either diabetes mellitus [36], post-splenectomy status [13] or haemolytic anaemia [6], we observed a sensitivity of 40% and a specificity of 75.5% for the mini-test versus our reference method. A near 20-fold lower cell/pore ratio appears as the single most important technical difference explaining the apparent lack of sensitivity of the mini-test. Suggestions are given as to how to modify the mini-test in order to obtain maximum sensitivity in the future.

A paediatric palliative care programme in development: trends in referral and location of death.

PURPOSE: To describe the formation of a paediatric palliative care programme providing care in hospital, at home or in hospice, ensuring continuity of care where the child and family desire. STUDY DESIGN: Descriptive analysis was performed on referral source, diagnosis and reason for discharge for patients referred to the Palliative Care Team at the Children's Hospital of Eastern Ontario in Ottawa, Ontario, Canada from 1999 to 2007. RESULTS: 341 children were referred. 24% had a neurological condition, 21% had genetic disorders or congenital anomalies, 20% had cancer, 18% had metabolic or neurodegenerative conditions and 17% had another diagnosis. The major sources of referral included paediatricians, neonatologists, oncologists and intensivists. 55% of the children have died. 58% of these died in hospital, 27% at home and 15% in hospice. Of the remaining 152 children, 7% were discharged from the programme after clinical improvement, 4% were moved to another geographic location or an adult centre, 2% were not eligible, 1% declined services and 4% were lost to follow-up. The remaining 90 children continue to be followed-up. In the hospitalised patients who died, the annual referral rate increased from 20% to >50%. IMPLICATIONS: Referral to the palliative care team has increased over time in all diagnostic categories and from all sources. Most children died in hospital; however, a significant number of families chose end-of-life care at home or in a hospice.

Talking with patients. Is it different when they are dying?

Talking with dying patients is different in some ways from talking with patients who do not have life-threatening illness; it can be frightening for physicians who have little experience. It is important to realize that, ultimately, the principles remain the same, and that the differences arise in the context and goals of the intervention. This article draws upon bedside experience to define several principles useful in talking with dying patients. Case examples illustrate each principle.

Latex-safe carts for bedside patient care.

The need to provide a latex-safe environment for patients with latex sensitivity continues to grow as the incidence of allergy increases. The unique approach of a mobile, latex-safe cart for use throughout the entire hospital is described, as well as the process used to develop the cart. A list of supplies for the cart is given to aid others who are faced with this task.

The measurement of erythrocyte deformability using micropore membranes. A sensitive technique with clinical applications.

This article describes a positive-pressure filtration technique using low pore-density, 3 microns pore-diameter polycarbonate membranes employed to evaluate erythrocyte deformability in a clinically oriented hematology laboratory. Mean erythrocyte resistance to filtration was expressed by a numerical index which takes into account both the initial resistance of dilute red cell suspensions to passage across the membrane and the relative pressure rise observed after filtration of 30 ml of the same suspension (3 . 10(8) cells). The resistance index of 99 blood samples obtained from 88 healthy adult volunteers ranged between 1.4 and 2.9, with normal Gaussian distribution and mean of 2.04. Values obtained on similarly prepared samples from 20 patients with various hemolytic anemias always fell outside this range, indicating a reduced deformability in every case. The existence of a strong positive correlation was found between the resistance index and the degree of reticulocytosis in these patients. This method appears more sensitive than previously described filtration techniques in detecting the presence of small numbers of poorly deformable erythrocytes in vivo, while being more practical and statistically more significant than the micropipette elastimetry technique. Our results, however, raise a new question concerning the role of reticulocytes in the evaluation of red cell deformability on blood samples from patients with hemolytic anemia.

Can we prevent an increase in the incidence of congenital rubella syndrome in the next decade?

The immunity to rubella of 115 girls aged 10 to 14 years was tested in 1978. The proportion of girls found to be immune was 80%, similar to rates in the prevaccination era. Nearly half of the immunity was from documented vaccination, and the other half was presumably from infection with wild rubella virus. The vaccination failure rate was 12%. Because of declining immunity to rubella of women of child-bearing age, detecting low levels of immunity in these women is becoming increasingly important. Immunization of 12- to 15-month-old children has not been effective. Vaccinating all girls 10 to 12 years old would likely be the most effective method of preventing an increase in the incidence of congenital rubella syndrome in the next decade.

Erythrocyte populations in pyruvate kinase deficiency anaemia following splenectomy. II. Cell deformability.

The deformability of three previously described morphological populations of erythrocytes from splenectomized pyruvate kinase (PK) deficient patients was assessed using both Nucleopore filtration and micropipette techniques. Mean deformability was decreased in fresh cells from all patients studied. Selective measurements indicated that immature lobulated reticulocytes in each patient were the least deformable cells, followed by bizarre echinocytes characterized by a hyperviscous behaviour, and by thin macrocytic discocytes whose values were closest to those of control erythrocytes. We conclude that poorly deformable reticulocytes in these patients, after initial trapping in the spleen, are unable to return to the circulation because of their inordinate dependence on mitochondrial oxydative phosphorylation for maintenance of a sufficient intracellular concentration of adenosine triphosphate (ATP); hence their rapid and selective destruction. Splenectomy therefore may be beneficial by allowing such cells to mature more slowly in the circulation of these patients until they develop crenation and hyperviscosity, two manifestations of profound ATP depletion possibly leading to destruction by the liver.