Quantitative measurement of IgE antibodies to purified allergens using streptavidin linked to a high-capacity solid phase

BACKGROUND: Commercially available assays for IgE antibody provide results in international units per milliliter for many allergen extracts, but this is not easily achieved with purified or novel allergens. OBJECTIVE: To develop assays for IgE antibody suitable for purified or novel allergens by using a commercially available immunosorbent. METHODS: Streptavidin coupled to a high-capacity immunosorbent (CAP) was used to bind biotinylated purified allergens from mite (Der p 1 and Der p 2), cat (Fel d 1), and dog (Can f 1). Assays for IgE antibody to these allergens were performed on sera from children (asthma and control) as well as adults with atopic dermatitis. RESULTS: The results were validated by serial dilution of sera with high and low levels of IgE antibody and were quantitated in international units per milliliter by using a standard curve. Values for IgE antibody to Der p 1, Der p 2, and Fel d 1 correlated with values obtained with the allergen extracts (r2 = 0.80, 0.84, and 0.95, respectively; P < .001 in each case). Furthermore, the values for IgE antibody in sera from children with high exposure to mite and cat allergens demonstrated 10-fold higher levels of IgE antibody to Der p 1 and Der p 2 than to Fel d 1 (P < .001). CONCLUSION: The streptavidin immunosorbent technique provides a new method for quantifying IgE antibody to purified proteins. The results provide evidence about the high quantities of IgE antibody to purified inhalant allergens in patients with atopic dermatitis. In addition, the results demonstrate major differences in IgE antibodies specific for mite and cat allergens among children with high exposure to both allergens.     

Severe dilated cardiomyopathy as a consequence of Ecstasy intake

Dilated cardiomyopathy (DCM) is one of the most common causes of heart failure with a prevalence of 1:2500. There are several primary and secondary etiologic factors, including gene mutations, infection agents, particularly viruses, toxins, autoimmune, and systemic disorders, and pheochromocytoma, neuromuscular, metabolic, mitochondrial, and nutritional disorders. However, a precise diagnosis can be reached only in no more than 50% of all cases. Herein, we report a rare case of hepatic damage and severe DCM as a consequence of relatively popular socially used narcotic-Ecstasy (3,4-methylenedioxy-N-methylamphetamine [MDMA]).     

Palmitic acid metabolism in the soleus muscle in vitro in hypo- and hyperthyroid rats

The aim of this study was to establish whether the rate of fatty acid (FA) incorporation and its utilization by the isolated soleus muscle is modified under conditions of thyroid hormone deficit or excess. The rate of palmitic acid (PA) uptake, oxidation and incorporation into intramuscular lipids with increasing PA concentration (0.5–1.5 mM) in the incubation medium were determined. In hypothyroid rats intramuscular triacylglycerol (TG) synthesis was increased, while the rate of PA oxidation to CO₂ and incorporation into mono- and diacylglycerols (MG/DG) and phospholipids (PL) remained unchanged. In rats with triiodothyronine (T₃) excess the rate of all processes studied was enhanced, although the percentage incorporation of PA into different classes of intramuscular lipids was fairly constant and, independently of thyroid state and FA concentration in the medium, was 56–66% for TG, 9-14% for MG/DG and 24–32% for PL. Our results thus indicate that even short-term T₃ excess accelerates the rate of FA uptake and metabolism in the oxidative soleus muscle, whereas in hypothyroid rats only intramuscular TG synthesis is affected.     

An immunoblotting procedure for screening glycophorins and band 3 protein in the same blots: Identification of glycophorin and band 3 variant forms

An immunoblotting procedure is described which makes it possible to screen multiple blood samples for the presence of glycophorin and band 3 variant forms with altered electrophoretic mobility. The procedure can be simplified by using whole red blood cell hemolysates instead of membranes for SDS-polyacrylamide gel electrophoresis. The use of hemolysates also has the advantage that antigens sensitive to proteolysis are not degraded in vitro. The same nitrocellulose blots were used for immunoenzymatic detection of glycophorins with a set of anti-glycophorin monoclonal antibodies, and for autoradiographic detection of band 3-derived bands with ¹²⁵I-labeled anti-band 3 monoclonal antibody. The screening of 157 Caucasian blood samples revealed the presence of a slower-migrating form of band 3 in seven cases and variant glycophorin in one case. The variant glycophorin exhibited the features of hybrid glycophorin of B-A type.     

The cGMP synthesis and PKG1 expression in murine lymphoid organs

Numerous reports indicate that cyclic 3',5' guanosine monophosphate (cGMP) is involved in the regulation of immune processes. However, the mechanisms responsible for the synthesis of this nucleotide and its signaling pathways in immune cells are still not well recognized. The aim of our studies was to establish: 1) which form of guanylyl cyclase (GC) synthesizes cGMP in murine lymphoid organs and 2) whether the same organs express the isoforms PKG1alpha and/or PKG1beta of protein kinase G, known as possible target for synthesized cGMP. Cells isolated from thymus, lymph nodes, and spleen were treated with activators (SNP, ANP, CNP, STa) of soluble or particulate cyclases. Sodium nitroprusside (SNP) elevated intracellular cGMP 2-fold in thymic and lymph node cells and about 10-fold in spleen cells. Atrial natriuretic peptide (ANP) caused modest but statistically significant increases of cGMP in cells of all three organs. Additionally, spleen cells elevated their cGMP content about 2-fold in response to C-type natriuretic protein (CNP). In cellular homogenates of the all analyzed organs, the antibody anti-PKG1beta stained the 78 kDa band corresponding to the molecular mass of PKG1. Only homogenates of spleen cells were stained by the antibody recognizing PKG1alpha. Our results indicate that in the investigated organs cGMP may be synthesized mainly by soluble GC in response to nitric oxide. The modest increase of cGMP upon stimulation by ANP suggests that in all these organs either exists only a small subpopulation of cells that express particulate cyclase GC-A or GC-A is expressed at very low level. In spleen cells, however, cyclase GC-B appears to be the more active enzyme. Elevated cGMP concentration may in turn activate PKG1beta in thymus, lymph node, and spleen cells and also PKG1alpha in spleen cells.     

Analgesic efficacy of sufentanil in dressings after surgical treatment of burn wounds

BackgroundThe aim of this study was to assess the analgesic efficacy of sufentanil in dressings after surgical treatment of burn wounds.Patients and methodsTwenty adult patients, who underwent surgical treatment of third–degree burn wounds under general anesthesia, were included. Two of the patients underwent surgery twice. During surgery, patients received 50–100 μg fentanyl every 20–30 min and, after surgery, patients received 100 mg ketoprofen twice daily. Additionally, ten patients (group 1) received 50 μg sufentanil added to the burn wound dressings soaked in octenidine and phenoxyethanol while 10 patients (group 2) received 25 μg sufentanil added to the same dressings. The rescue analgesic, which was administered when pain intensified, was 5 mg subcutaneous morphine. Plasma sufentanil concentrations were assayed at 1, 2, 3, and 6 h after surgery completion and when pain was reported, along with pain intensity evaluation.ResultsSufentanil was not detected in the serum of any patients. Rescue morphine was given during the postoperative period (24 h) in one patient in group 1 (who underwent surgery twice) and three patients in group 2. The mean sufentanil concentration in dressings was higher in group 1 (0.13 ± 0.03) than group 2 (0.06 ± 0.03 μg/mL; p < 0.001). The group 1 patient who received rescue morphine had a sufentanil concentration of 0.10 μg/mL, which was the lowest concentration in group 1. Group 2 patients who received rescue morphine had sufentanil concentrations of at least two–fold lower (0.03–0.05 μg/mL). No adverse effects were observed.ConclusionsSufentanil in dressings after burn wound surgery provides effective and safe analgesia and the sufentanil concentration in dressings should be ≥0.10 μg/mL in a solution of octenidine and phenoxyethanol.