Buffy Coat Smears Research Articles

Biosynthesis of von Willebrand protein by human megakaryocytes.

Immunofluorescence staining of buffy coat smears from a patient with chronic myelogenous leukemia in accelerated phase showed that approximately 13% of all nucleated cells contained von Willebrand protein and, therefore, appeared to be of megakaryocytic origin. This was confirmed by positive staining with antisera against platelet factor 4 and platelet glycoproteins. Short-term cultures of the buffy coat, which lacked endothelial cells, were metabolically labeled with [35S]methionine, and von Willebrand protein was immunopurified from cell lysates and culture medium. Cultures from this patient synthesized and secreted von Willebrand protein, in contrast with cultures from other patients with leukemia, who lacked circulating megakaryocytes, and from normal volunteers. The subunit composition of the megakaryocytic von Willebrand protein was very similar to that of human umbilical vein endothelial cells. The size of the processed subunit (220 kD) and of the cellular (260 kD) and secreted (275 kD) precursors from the two cell types were indistinguishable by gel electrophoresis. Furthermore, the ratio of precursor to processed subunit and the pattern of cellular and secreted nonreduced multimers were very similar. It appears, therefore, that the processing steps in biosynthesis of von Willebrand protein used by the megakaryocytes are very similar to those of umbilical vein endothelial cells.

Buffy coat smears of blood drawn through central venous catheters as an aid to rapid diagnosis of systemic fungal infections

Buffy coat smears of blood drawn through central venous catheters as an aid to rapid diagnosis of systemic fungal infections

Rapid diagnosis of neonatal bacteremia with acridine organe-stained buffy coat smears

Rapid diagnosis of neonatal bacteremia with acridine organe-stained buffy coat smears

Use of buffy-coat smears in the diagnosis of septicaemia in children.

Gram-stained buffy-coat smears from venous and capillary blood samples in 105 children suspected of septicaemia were examined for the presence of bacteria and the results compared with blood culture isolates. Gram-positive and Gram-negative bacteria were identified in 18 venous (44%) and 19 capillary (46%) buffy-coat preparations in 41 instances where bacterial organisms were isolated from the blood cultures. It is concluded that the examination of buffy-coat smears for bacteria in children suspected of septicaemia is a useful adjunct to blood cultures and, in areas where no facilities exist for culture of blood, may be a simple and rapid method of establishing the diagnosis of bacteraemia in suspected patients.

The leukocyte composition of peripheral blood buffy coat.

To assist in the interpretation of buffy coat smears from leukopenic patients, we determined the composition of the buffy coat of the blood of normal volunteers. Leukocytes were concentrated by centrifugation in microhematocrit tubes. Stained smears of these cells showed minor differences in the proportions of neutrophils, lymphocytes, and monocytes when compared to unconcentrated peripheral blood. Granulocytes of greater immaturity than the metamyelocyte were not encountered, and their presence should suggest the possibility of marrow dysfunction. Occasional young monocytoid cells and "stimulated" lymphocytes were seen.

Fungemia in chronic cavitary pulmonary histoplasmosis.

Syndrome. A 76-year-old man with chronic obstructive pulmonary disease developed a progressive cavitary lesion of the right-upper lobe. Studies for neoplasm and acid-fast bacilli were negative. Diagnosis. Buffy-coat smears were prepared by the capillary method for leukocyte concentrates [1] and treated with Wright-Giemsa stain. The smears showed small (2-3 Am) organisms containing dark refractile bodies (figure). Most of the organisms present were extracellular. Cultures of the blood and sputum were later found to be positive for Histoplasma capsul-

Preparation of Buffy Coat Smears in Leukopenic Patients

Although clinically important, white cell differential counts cannot be accurately performed on peripheral blood smears of severely leukopenic patients. A simple technique is described for preparation of buffy coat smears from leukopenic blood that preserves white cell morphology and gives a reasonably accurate representation of the white cell differential count.

Evaluation of the Buffy-Coat Smear for Rapid Detection of Bacteremia

Buffy-coat smears prepared from 599 blood samples, obtained simultaneously with blood for culture, were examined for bacteria. Each blood sample was quantitatively cultured on chocolate agar. Twenty-one blood cultures (3.6%) from 17 patients were positive. Two (11.7%) of the 17 patients had positive buffy-coat smears. One patient had two positive smears, for a total of three positive buffy-coat smears from 21 positive blood cultures. Chart reviews were performed for two patients with positive buffy-coat smears, three patients with positive blood cultures (containing the same organisms as the two study patients) but negative buffy-coat smears, and six additional patients who had had positive buffy-coat smears between 1974 and 1979. Of the eight patients with positive smears, six (75%) were infected with Gram-positive cocci, three of which were Streptococcus pneumoniae. Mortality was 100%.

Evaluation of the buffy-coat smear for rapid detection of bacteremia

Evaluation of the buffy-coat smear for rapid detection of bacteremia

Rapid method for determining T lymphocyte levels using acid alpha-naphthyl acetate esterase.

A method for rapidly determining levels of T lymphocytes in humans is presented. The method involves staining lymphocytes in peripheral blood or buffy coat smears for acid alpha-naphthylacetate esterase for 1 hr at 37 C. After counterstaining with Wright's stain, the percentage of esterase-positive lymphocytes accurately reflects the percentage of lymphocytes forming E rosettes with sheep erythrocytes.

Acid alpha-naphthyl acetate esterase activity in stored buffy coat smears.

The stability of acid alpha-naphthyl acetate esterase activity in smears of buffy coats was studied. Fixed smears may be stored up to 20 days at 25 degrees C. Unfixed smears may be stored up to 24 hr at 25 degrees C before there is a significant decrease in activity. In constant, fixed or unfixed smears held at -10 degrees C begin to lose activity within 24 hr.

802 CAN NEONATAL SEPSIS BE DIAGNOSED EARLY?

Currently, many babies receive antibiotics unnecessarily for “phantom” infections. One-third of babies (258/800) admitted to our intensive care nursery in approximately 2 years had bacterial cultures of blood, urine and CSF, and received antibiotics. Several tests which can be rapidly and easily performed were combined to predict neonatal infection. These included WBC and diff., “mini”-sed. rate, latex C-reactive protein, haptoglobin and IgM. The scoring system previously described (Pediatr. Res. 11:504, 1977) was used. Of 20 cases of proven infection, 17 (85%) had elevated scores (the other 3 babies died very rapidly). In 32 babies with possible or probable infection (antibiotics given for 5 or more days without positive blood culture), 23 (72%) had an elevated score. In 206 babies without infection (antibiotics for 3 or less days), only 8 (4%) had an elevated score. Individual tests were predictive in 33% or less of proven cases of infection. Substituting buffy-coat smears for latex-IgM could improve the predictive value of this scoring system. These easily performed tests require no special laboratory facilities and provide an accurate diagnostic indication of neonatal sepsis within 1 hour. This could save physician's time and decrease antibiotic use, length of hospital stay and nosocomial infection.

Early diagnosis of neonatal bacteremia by buffy-coat examination.

This study demonstrates the value of the buffy-coat smear examination in the early diagnosis of neonatal bacteremia. It is inexpensive, easy to perform, and relatively efficient. In most situations large numbers of bacteria could be seen in each positive smear. Unfortunately, the presence of a positive buffy-coat smear was often associated with a poor prognosis; more than one half (four of seven) of infants with positive smears died.

An autoradiographic study of human blood basophils.

The aim of this paper is the selective visualization of human blood basophils in autoradiographs. [(3)H]Thymidine-labelled basophils in buffy coat smears were fixed in methanol-formaldehyde follwed by 5-aminoacridine hydrochloride and stained with basic Aldehyde Fuchsin prior to autoradiographic processing. The technique described represents a simple method for quantitative autoradiographic studies on basophil kinetics and the interaction of these cells with IgE-mediated atopias.

Demonstration of Neisseria gonorrhoeae with fluorescent antibody in patients with disseminated gonococcal infection.

Among patients with disseminated gonococcal infection and pustular skin lesions, Neisseria gonorrhoeae was identified in exudates from the skin lesions by the direct fluorescent antibody test in 57%, by gram stain in 11%, and by culture in only 9%. However, gonococci were cultured from four of seven pustular biopsies of skin lesions. Gonococci were also identified by fluorescent antibody examination in four of eight buffy coat smears from patients with negative blood cultures, but in only one of eight culture-negative synovial fluids. Among patients without disseminated gonococcal infection, smears of synovial fluid, buffy coat, and exudate from skin pustules were uniformly negative by the fluorescent antibody test. This test has contributed to studies of the pathogenesis of disseminated gonococcal infection and is frequently useful in the diagnosis of this condition.

GIANT NUCLEAR MASSES IN THE LUNGS AND BLOOD IN MALIGNANT MALARIA

Numerous giant nuclear masses were seen in sections of lung from two fatal cases of subtertian malaria. Similar structures were found in all of ten buffy-coat smears and thirty thick films of the blood of one of these cases taken 7 hours before death. It is suggested that the detection of these nuclear structures in thick bloodfilms, buffy-coat smears, or after more elaborate concentration techniques may prove to be a valuable indication of disseminated intravascular coagulation in malignant malaria.

Early Diagnosis of Bacteremia by Buffy-Coat Examinations

A smear of leukocytes from the buffy-coat layer was obtained when peripheral blood specimens were taken for culture. Fourteen of 135 blood cultures were positive, and five of the 14, from three patients, showed Gram-positive intracellular organisms on Gram stain of the buffy-coat layer. Grampositive cocci were seen in two of three patients with staphylococcal bacteremia and Gram-positive rods in one patient with<i>Clostridium perfringens</i>bacteremia. These two cases illustrate the diagnostic role of the buffy-coat smear and its use in the early diagnosis of bacteremia.

Diagnosis of Tropical Canine Pancytopaenia (Ehrlichia canis Infection) by Immunofluorescence

SUMMARY Ehrlichia canis was demonstrated by direct immunofluorescence in buffy coat and other tissue smears using fluorescein conjugates prepared from the sera of infected dogs. The antigenic forms of E. canis demonstrated by immunofluorescence were identical with those demonstrated by Giemsa’s stain. Low magnification scanning of immunofluorescent preparations was quicker and more sensitive than the examination of similar preparations stained with Giemsa. E. canis could also be demonstrated by staining with acridine orange. Tests for infcctivity in dogs were more sensitive than any of these 3 visual methods. Sera and smears from dogs infected with strains of E. canis isolated from widely separated areas of the world were examined by immunofluorescence and all appeared to be identical antigenically.

The localization of Australia antigen by immunofluorescence.

We have studied the localization of Australia antigen, a particulate substance associated with hepatitis, by means of the fluorescent antibody technique. Preparations were made from 61 liver biopsy specimens taken from patients with infectious hepatitis, serum hepatitis, and a variety of other diseases. When tested with fluorescein-conjugated rabbit anti-Au(1) antisera all 26 patients who had Au(1) in their serum had specific fluorescence in their liver cells. The fluorescence appeared in three forms: as discrete particles within the nucleus, diffuse fluorescence of the entire nucleus, and fluorescence of the nuclear rim. Occasionally there were also fluorescent particles in the cytoplasm. Other specimens were tested with the fluorescent antibody including a variety of human tissues, buffy coat smears, peripheral lymphocyte cultures, and cells obtained from bile and duodenal drainage. Among these specimens, fluorescence was found in the cytoplasm of a few cells in the bone marrow of two patients with hepatitis and Au(1) in their serum, and in the liver, spleen, mesentery, and testis of one patient with leukemia, chronic hepatitis, and Au(1) in his serum. We have shown that the presence of fluorescent particles in the liver cells is strongly associated with the presence of Au(1) in the serum and the diagnosis of viral hepatitis. We believe that this study adds support to the hypothesis that Australia antigen is an antigenic determinant of a virus capable of causing hepatitis.

HISTOCHEMICAL DEMONSTRATION OF SULFATED MUCOSUBSTANCES AND CATIONIC PROTEINS IN HUMAN GRANULOCYTES AND PLATELETS

Histochemical staining methods visualize sulfated mucosubstance in numerous small granules of early neutrophil leukocytes in appropriately processed human bone marrow smears. These reactive granules presumably constitute the counterpart in human neutrophils of the mucosaccharide-rich primary granules in rabbit heterophils. Neutrophils at a late developmental stage in human marrow or buffy coat smears reveal few or no granules with such reactivity. Strong staining for sulfated mucosubstance is readily demonstrable in numerous large cytoplasmic granules in human eosinophils. This reactivity diminishes but does not disappear during maturation of the eosinophil and its single population of cytoplasmic granules. Granules of human basophils at all developmental stages stain for sulfated mucosubstance. Platelets in human buffy coat smears reveal evidence for sulfated mucosaccharide. The mucosaccharide-containing granules of the three myeloid series also disclose acidophilia at high pH indicative of the presence of strongly cationic proteins.