Measuring Electrical And Mechanical Properties Of Red Blood Cells With A Double Optical Tweezers

The fluid lipid bilayer viscoelastic membrane of red blood cells (RBC) contains antigen glycolproteins and proteins which can interact with antibodies to cause cell agglutination. This is the basis of most of the immunohematologic tests in blood banks and the identification of the antibodies against the erythrocyte antigens is of fundamental importance for transfusional routines. The negative charges of the RBCs creates a repulsive electric (zeta) potential between the cells and prevents their aggregation in the blood stream. The first counterions cloud strongly binded moving together with the RBC is called the compact layer. This report proposes the use of a double optical tweezers for a new procedure for measuring: (1) the apparent membrane viscosity, (2) the cell adhesion, (3) the zeta potential and (4) the compact layer's size of the charges formed around the cell in the electrolytic solution. To measure the membrane viscosity we trapped silica beads strongly attached to agglutinated RBCs and measured the force to slide one RBC over the other as a function of the relative velocity. The RBC adhesion was measured by slowly displacing two RBCs apart until the disagglutination happens. The compact layer's size was measured using the force on the silica bead attached to a single RBC in response to an applied voltage and the zeta potential was obtained by measuring the terminal velocity after releasing the RBC from the optical trap at the last applied voltage. We believe that the methodology here proposed can improve the methods of diagnosis in blood banks.6326Eylar, E.H., Madoff, M.A., Brody, O.V., Oncley, J.L., The contribution of sialic acid to the surface charge of the erythrocyte (1962) J. Biol. Chem., 237, pp. 1992-2000Pollack, W., Reckel, R.P., A reappraisal of the forces involved in Hemagglutination (1977) Int Archs Allergy Appl. Immun., 54, pp. 29-42Ashkin, A., Dziedzic, J.M., Bjorkholm, J.E., Chu, S., Observation of a single-beam gradient force trap for dielectric particles (1986) Opt. Lett., 11, pp. 288-290Ashkin, A., Dziedzic, J.M., Optical trapping and manipulation of viruses and bacteria (1987) Science, 235, pp. 1517-1520Grier, D.G., A revolution in optical manipulation (2003) Nature, 424, pp. 810-816Zhu, C., Bao, G., Wang, N., Cell Mechanics: Mechanical response, cell adhesion, and molecular deformation (2000) Annu. Rev. Biomed. Eng., 2, pp. 189-226Neuman, K.C., Block, S., Optical trapping (2004) Rev. Sci. Instrum., 75, pp. 2787-2809Saffman, P.G., Delbruck, M., Brownian motion in biological membranes (1975) Proc. Nat. Acad. Sci. USA, 72, pp. 3111-3113Dimova, R., Danov, K., Pouligny, B., Ivanov, I.B., Drag of a solid particle trapped in a thin film or at an interface: Influence of surface viscosity and elasticity (2000) J. Colloid and Interface Science, 226, pp. 35-43Hochmuth, R., Worthy, P., Evans, E., Red cell extensional recovery and the determination of membrane viscosity (1979) Biophys. J., 26, pp. 101-114Sze, A., Erickson, D., Ren, L., Li, D., Zeta-potential measurement using the Smoluchowski equation and the slope of the current-time relationship in electroosmotic flow (2003) J. Colloid and Interface Science, 261, pp. 402-410Hunter, R.J., (1981) Zeta Potential in Colloid Science, , Academic Press, New YorkPollack, W., Hager, H.J., Reckel, R., Toren, D.A., Singher, H.O., A study of the forces involved in the second stage of hemaggltination (1965) Transfusion, 5, pp. 158-183Chelidze, T., Dielectric spectroscopy of blood (2002) J. Non-crystalline Solids, 305, pp. 285-294Hymer, W.C., Barlow, G.H., Blaisdell, S.J., Continuous flow electrophoretic separation of proteins and cells from mammalian tissues (1987) Cell Biophys., 10, pp. 61-85Hashimoto, N., Fujita, S., Yokoyama, T., Cell electrophoretic mobility and glycerol lysis of human erythrocytes in various diseases (1998) Electrophoresis, 19, pp. 1227-123

Mechanical Properties Of Stored Red Blood Cells Using Optical Tweezers

We have developed a method for measuring the red blood cell (RBC) membrane overall elasticity μ by measuring the deformation of the cells when dragged at a constant velocity through a plasma fluid by an optical tweezers. The deformability of erythrocytes is a critical determinant of blood flow in the microcirculation. We tested our method and hydrodynamic models, which included the presence of two walls, by measuring the RBC deformation as a function of drag velocity and of the distance to the walls. The capability and sensitivity of this method can be evaluated by its application to a variety of studies, such as, the measurement of RBC elasticity of sickle cell anemia patients comparing homozygous (HbSS), including patients taking hydroxyrea (HU) and heterozygous (HbAS) with normal donors and the RBC elasticity measurement of gamma irradiated stored blood for transfusion to immunosupressed patients as a function of time and dose. These studies show that the technique has the sensitivity to discriminate heterozygous and homozygous sickle cell anemia patients from normal donors and even follow the course of HU treatment of Homozygous patients. The gamma irradiation studies show that there is no significant change in RBC elasticity over time for up to 14 days of storage, regardless of whether the unit was irradiated or not, but there was a huge change in the measured elasticity for the RBC units stored for more than 21 days after irradiation. These finds are important for the assessment of stored irradiated RBC viability for transfusion purposes because the present protocol consider 28 storage days after irradiation as the limit for the RBC usage.593016Ashkin, A., Dziedzic, J.M., Optical trapping and manipulation of viruses and bacteria (1987) Science, 235, pp. 1517-1520Barjas-Castro, M.L., Brandão, M.M., Fontes, A., Costa, F.F., Cesar, C.L., Saad, S.T.O., Elastic properties of irradiated red blood cell units measured by optical tweezer (2002) Transfusion, 42, pp. 1196-1199Brandão, M.M., Fontes, A., Barjas-Castro, M.L., Barbosa, L.C., Costa, F.F., Cesar, C.L., Saad, S.T.O., Optical tweezers for measuring red blood cell elasticity: Application to the study of drug response in sickle cell disease (2003) European Journal of Haematology, 70, pp. 207-211Williamson, L.M., Warwick, R.M., Transfusion-associated graft-versus-host disease and its prevention (1995) Blood Rev., 9, pp. 251-261Button, L.N., Dewolf, W.C., Newburger, P.E., The effecr of irradiation on blood components (1981) Transfusion, 21, pp. 419-426Platt, O.S., The sickle syndrome (1995) Blood: Principles and Practice of Hematology, , R. I Hadlin, S. E. Lux, T. P. Stossel, J. B. Lippincott, PhiladelphiaBallas, S.K., Dover, G.J., Charache, S., Effect of hydroxyurea on the rheological properties of sickle erythrocytes in vivo (1989) Am. J. Hematol, 32, pp. 104-111Groner, W., Mohandas, N., Bessis, M., New optical technique for measuring erythrocyte deformability with the ektacytometer (1980) Clin. Chem., 26, pp. 1435-1442De Franceschi, L., Bachir, D., Galacteros, F., Tchernia, G., Cynober, T., Alper, S., Platt, O., Brugnara, C., Oral magnesium supplements reduce erythrocyte dehydration in patients with sickle cell disease (1997) J Clin Invest, 100, pp. 1847-1852Hochmuth, R.M., Worthy, P.R., Evans, E.A., Red cell extensional recovery and the determination of membrane viscosity (1979) Biophys. J., 26, pp. 101-114Evans, E.A., La Celle, P.L., Intrinsic material properties of the erythrocyte membrane indicated by mechanical analysis of deformation (1975) Blood, 45, pp. 29-43Itoh, T., Chien, S., Usami, S., Effects of hemoglobin concentration on deformability of individual sickle cells after deoxygenation (1995) Blood, 85, pp. 2245-2253Evans, E.A., Mohandas, N., Membrane-associated sickle hemoglobin: A major determinant of sickle erythrocyte rigidity (1987) Blood, 70, pp. 1443-1449Dong, C., Chadwick, R.S., Schechter, A.N., Influence of sickle hemoglobin polymerization and membrane properties on deformability of sickle erythrocytes in the microcirculation (1992) Biophys. J., 63, pp. 774-783Suzuki, Y., Tateishi, N., Cicha, I., Decreased deformability of the X-ray irradiated red blood cells stored in manitol-adenine-phosphate medium (2000) Clin. Hemorheol. Micro-cire., 22, pp. 131-14

Studying Red Blood Cell Agglutination By Measuring Membrane Viscosity With Optical Tweezers

The red blood cell (RBC) viscoelastic membrane contains proteins and glycoproteins embedded in a fluid lipid bilayer that are responsible for cell agglutination. Manipulating RBCs rouleaux with a double optical tweezers, we observed that the cells slide easily one over the others but are strongly connected by their edges. An explanation for this behavior could be the fact that when the cells slide one over the others, proteins are dragged through the membrane. It confers to the movement a viscous characteristic that is dependent of the velocity between the RBCs and justifies why is so easy to slide them apart. Therefore, in a first step of this work, by measuring the force as a function of the relative velocity between two cells, we confirmed this assumption and used this viscous characteristic of the RBC rouleaux to determine the apparent membrane viscosity of the cell. As this behavior is related to the proteins interactions, we can use the apparent membrane viscosity to obtain a better understanding about cell agglutination. Methods related to cell agglutination induced by antigen-antibody interactions are the basis of most of tests used in transfusion centers. Then, in a second step of this work, we measured the apparent membrane viscosity using antibodies. We observed that this methodology is sensitive to different kinds of bindings between RBCs. Better comprehension of the forces and bindings between RBCs could improve the sensibility and specificity of the hemagglutination reactions and also guides the development of new potentiator substances.6644Fontes, A., Fernandes, H.P., Barjas-Castro, M.L., Thomaz, A.A., Pozzo, L., Barbosa, L.C., Cesar, C.L., Red blood cell membrane viscoelasticity, agglutination and zeta potential measurements with double optical tweezers (2006) Proceedings of SPIE, 6088, pp. 296-305Eylar, E.H., Madoff, M.A., Brody, O.V., Oncley, J.L., The contribution of sialic acid to the surface charge of the erythrocyte (1962) J. Biol. Chem, 237, pp. 1992-2000Pollack, W., Reckel, R.P., A reappraisal of the forces involved in Hemagglutination (1977) Int Archs Allergy Appl. Immun, 54, pp. 29-42Fontes, A., Giorgio, S., de Castro Jr., A.B., Neto, V.M., Pozzo, L.Y., Marques, G.P., Barbosa, L.C., Cesar, C.L., Determination of femto Newton forces and fluid viscosity using optical tweezers: Application to Leishmania amazonensis (2005) Proceedings of SPIE, 5699, pp. 419-425Saffman, P.G., Delbruck, M., Brownian motion in biological membranes (1975) Proc. Nat. Acad. Sci. USA, 72, pp. 3111-3113Dimova, R., Danov, K., Pouligny, B., Ivanov, I.B., Drag of a solid particle trapped in a thin film or at an interface: Influence of surface viscosity and elasticity (2000) J. Colloid and Interface Science, 226, pp. 35-4

Bartonella Clarridgeiae Bacteremia Detected In An Asymptomatic Blood Donor

Human exposure to Bartonella clarridgeiae has been reported only on the basis of antibody detection. We report for the first time an asymptomatic human blood donor infected with B. clarridgeiae, as documented by enrichment blood culture, PCR, and DNA sequencing.531352356Maggi, R.G., Duncan, A.W., Breitschwerdt, E.B., Novel chemically modified liquid medium that will support the growth of seven Bartonella species (2005) J Clin Microbiol, 43, pp. 2651-2655. , http://dx.doi.org/10.1128/JCM.43.6.2651-2655.2005Drummond, M.R., Pitassi, L.H., Lania, B.G., Dos Santos, S.R., Gilioli, R., Velho, P.E., Detection of Bartonella henselae in defibrinated sheep blood used for culture media supplementation (2011) Braz J Microbiol, 42, pp. 430-432. , http://dx.doi.org/10.1590/S1517-83822011000200003Altschul, S.F., Gish, W., Miller, W., Myers, E.W., Lipman, D.J., Basic local alignment search tool (1990) J Mol Biol, 215, pp. 403-410Dalton, M.J., Robinson, L.E., Cooper, J., Regnery, R.L., Olson, J.G., Childs, J.E., Use of Bartonella antigens for serologic diagnosis of cat-scratch disease at a national referral center (1995) Arch Intern Med, 155, pp. 1670-1676Breitschwerdt, E.B., Maggi, R.G., Chomel, B.B., Lappin, M.R., Bartonellosis: An emerging infectious disease of zoonotic importance to animals and human beings (2010) J Vet Emerg Crit Care (San Antonio), 20, pp. 8-30. , http://dx.doi.org/10.1111/j.1476-4431.2009.00496.xChamberlin, J., Laughlin, L.W., Romero, S., Solorzano, N., Gordon, S., Andre, R.G., Pachas, P., Watts, D., Epidemiology of endemic Bartonella bacilliformis: A prospective cohort study in a Peruvian mountain valley community (2002) J Infect Dis, 186, pp. 983-990. , http://dx.doi.org/10.1086/344054Maggi, R.G., Ericson, M., Mascarelli, P.E., Bradley, J.M., Breitschwerdt, E.B., Bartonella henselae bacteremia in a mother and son potentially associated with tick exposure (2013) Parasit Vectors, 6, p. 101. , http://dx.doi.org/10.1186/1756-3305-6-101Scott, M.A., McCurley, T.L., Vnencak-Jones, C.L., Hager, C., McCoy, J.A., Anderson, B., Collins, R.D., Edwards, K.M., Cat scratch disease: Detection of Bartonella henselae DNA in archival biopsies from patients with clinically, serologically, and histologically defined disease (1996) Am J Pathol, 149, pp. 2161-2167Slater, L.N., Welch, D.F., Min, K.W., Rochalimaea henselae causes bacillary angiomatosis and peliosis hepatis (1992) Arch Intern Med, 152, pp. 602-606Sander, A., Zagrosek, A., Bredt, W., Schiltz, E., Piemont, Y., Lanz, C., Dehio, C., Characterization of Bartonella clarridgeiae flagellin (FlaA) and detection of antiflagellin antibodies in patients with lymphadenopathy (2000) J Clin Microbiol, 38, pp. 2943-2948Kordick, D.L., Hilyard, E.J., Hadfield, T.L., Wilson, K.H., Steigerwalt, A.G., Brenner, D.J., Breitschwerdt, E.B., Bartonella clarridgeiae, a newly recognized zoonotic pathogen causing inoculation papules, fever, and lymphadenopathy (cat scratch disease) (1997) J Clin Microbiol, 35, pp. 1813-1818Margileth, A.M., Baehren, D.F., Chest-wall abscess due to cat-scratch disease (CSD) in an adult with antibodies to Bartonella clarridgeiae: Case report and review of the thoracopulmonary manifestations of CSD (1998) Clin Infect Dis, 27, pp. 353-357. , http://dx.doi.org/10.1086/514671Chomel, B.B., Mac Donald, K.A., Kasten, R.W., Chang, C.C., Wey, A.C., Foley, J.E., Thomas, W.P., Kittleson, M.D., Aortic valve endocarditis in a dog due to Bartonella clarridgeiae (2001) J Clin Microbiol, 39, pp. 3548-3554. , http://dx.doi.org/10.1128/JCM.39.10.3548-3554.2001Gillespie, T.N., Washabau, R.J., Goldschmidt, M.H., Cullen, J.M., Rogala, A.R., Breitschwerdt, E.B., Detection of Bartonella henselae and Bartonella clarridgeiae DNA in hepatic specimens from two dogs with hepatic disease (2003) J Am Vet Med Assoc, 222, pp. 47-51. , http://dx.doi.org/10.2460/javma.2003.222.47, 35Robinson, M.T., Hillman, T., Langton, D.A., Shaw, S.E., Bartonella clarridgeiae in a cat in the UK (2009) Vet Rec, 164, pp. 58-59. , http://dx.doi.org/10.1136/vr.164.2.58Sykes, J.E., Westropp, J.L., Kasten, R.W., Chomel, B.B., Association between Bartonella species infection and disease in pet cats as determined using serology and culture (2010) J Feline Med Surg, 12, pp. 631-636. , http://dx.doi.org/10.1016/j.jfms.2010.04.003Fouch, B., Coventry, S., A case of fatal disseminated Bartonella henselae infection (cat-scratch disease) with encephalitis (2007) Arch Pathol Lab Med, 131, pp. 1591-1594Boudebouch, N., Sarih, M., Beaucournu, J.C., Amarouch, H., Hassar, M., Raoult, D., Parola, P., Bartonella clarridgeiae, B. Henselae, and Rickettsia felis in fleas from Morocco (2011) Ann Trop Med Parasitol, 105, pp. 493-498. , http://dx.doi.org/10.1179/1364859411Y.0000000038Kordick, D.L., Brown, T.T., Shin, K., Breitschwerdt, E.B., Clinical and pathologic evaluation of chronic Bartonella henselae or Bartonella clarridgeiae infection in cats (1999) J Clin Microbiol, 37, pp. 1536-1547Chomel, B.B., Carlos, E.T., Kasten, R.W., Yamamoto, K., Chang, C.C., Carlos, R.S., Abenes, M.V., Pajares, C.M., Bartonella henselae and Bartonella clarridgeiae infection in domestic cats from the Philippines (1999) Am J Trop Med Hyg, 60, pp. 593-597Dehio, C., Bartonella interactions with endothelial cells and erythrocytes (2001) Trends Microbiol, 9, pp. 279-285. , http://dx.doi.org/10.1016/S0966-842X(01)02047-9Dehio, C., Meyer, M., Berger, J., Schwarz, H., Lanz, C., Interaction of Bartonella henselae with endothelial cells results in bacterial aggregation on the cell surface and the subsequent engulfment and internalisation of the bacterial aggregate by a unique structure, the invasome (1997) J Cell Sci, 110 (18), pp. 2141-2154Braga Mdo, S., Diniz, P.P., André, M.R., Bortoli, C.P., Machado, R.Z., Molecular characterisation of Bartonella species in cats from São Luís, state of Maranhão, North-Eastern Brazil (2012) Mem Inst Oswaldo Cruz, 107, pp. 772-777. , http://dx.doi.org/10.1590/S0074-02762012000600011Eremeeva, M.E., Gerns, H.L., Lydy, S.L., Goo, J.S., Ryan, E.T., Mathew, S.S., Ferraro, M.J., Koehler, J.E., Bacteremia, fever, and splenomegaly caused by a newly recognized Bartonella species (2007) N Engl J Med, 356, pp. 2381-2387. , http://dx.doi.org/10.1056/NEJMoa065987Chomel, B.B., Boulouis, H.J., Breitschwerdt, E.B., Kasten, R.W., Vayssier-Taussat, M., Birtles, R.J., Koehler, J.E., Dehio, C., Ecological fitness and strategies of adaptation of Bartonella species to their hosts and vectors (2009) Vet Res, 40, p. 29. , http://dx.doi.org/10.1051/vetres/2009011Breitschwerdt, E.B., Maggi, R.G., Duncan, A.W., Nicholson, W.L., Hegarty, B.C., Woods, C.W., Bartonella species in blood of immunocompetent persons with animal and arthropod contact (2007) Emerg Infect Dis, 13, pp. 938-941. , http://dx.doi.org/10.3201/eid1306.061337Carson, J.L., Grossman, B.J., Kleinman, S., Tinmouth, A.T., Marques, M.B., Fung, M.K., Holcomb, J.B., Djulbegovic, B., Red blood cell transfusion: A clinical practice guideline from the AABB (2012) Ann Intern Med, 157, pp. 49-58. , http://dx.doi.org/10.7326/0003-4819-157-1-201206190-00429Ramirez-Arcos, S., Goldman, M., Blajchman, M., Bacterial contamination (2012) Transfusion Reaction, 4, pp. 153-189. , Popovsky MA (ed), American Association Of Blood Banks, Bethesda, MDVamvakas, E.C., Blajchman, M.A., Transfusion-related mortality: The ongoing risks of allogeneic blood transfusion and the available strategies for their prevention (2009) Blood, 113, pp. 3406-3417. , http://dx.doi.org/10.1182/blood-2008-10-167643Magalhães, R.F., Cintra, M.L., Barjas-Castro, M.L., Del Negro, G.M., Okay, T.S., Velho, P.E., Blood donor infected with Bartonella henselae (2010) Transfus Med, 20, pp. 280-282. , http://dx.doi.org/10.1111/j.1365-3148.2010.01001.xMagalhães, R.F., Pitassi, L.H., Salvadego, M., De Moraes, A.M., Barjas-Castro, M.L., Velho, P.E., Bartonella henselae survives after the storage period of red blood cell units: Is it transmissible by transfusion? (2008) Transfus Med, 18, pp. 287-291. , http://dx.doi.org/10.1111/j.1365-3148.2008.00871.xLin, J.W., Chen, C.M., Chang, C.C., Unknown fever and back pain caused by Bartonella henselae in a veterinarian after a needle puncture: A case report and literature review (2011) Vector Borne Zoonotic Dis, 11, pp. 589-591. , http://dx.doi.org/10.1089/vbz.2009.0217Oliveira, A.M., Maggi, R.G., Woods, C.W., Breitschwerdt, E.B., Suspected needle stick transmission of Bartonella vinsonii subspecies berkhoffii to a veterinarian (2010) J Vet Intern Med, 24, pp. 1229-1232. , http://dx.doi.org/10.1111/j.1939-1676.2010.0563.xOhl, M.E., Spach, D.H., Bartonella quintana and urban trench fever (2000) Clin Infect Dis, 31, pp. 131-135. , http://dx.doi.org/10.1086/313890Daly, J.S., Worthington, M.G., Brenner, D.J., Moss, C.W., Hollis, D.G., Weyant, R.S., Steigerwalt, A.G., O'Connor, S.P., Rochalimaea elizabethae sp. Nov. Isolated from a patient with endocarditis (1993) J Clin Microbiol, 31, pp. 872-881Oksi, J., Rantala, S., Kilpinen, S., Silvennoinen, R., Vornanen, M., Veikkolainen, V., Eerola, E., Pulliainen, A.T., Cat scratch disease caused by Bartonella grahamii in an immunocompromised patient (2013) J Clin Microbiol, 51, pp. 2781-2784. , http://dx.doi.org/10.1128/JCM.00910-13Breitschwerdt, E.B., Mascarelli, P.E., Schweickert, L.A., Maggi, R.G., Hegarty, B.C., Bradley, J.M., Woods, C.W., Hallucinations, sensory neuropathy, and peripheral visual deficits in a young woman infected with Bartonella koehlerae (2011) J Clin Microbiol, 49, pp. 3415-3417. , http://dx.doi.org/10.1128/JCM.00833-11Raoult, D., Roblot, F., Rolain, J.M., Besnier, J.M., Loulergue, J., Bastides, F., Choutet, P., First isolation of Bartonella alsatica from a valve of a patient with endocarditis (2006) J Clin Microbiol, 44, pp. 278-279. , http://dx.doi.org/10.1128/JCM.44.1.278-279.2006Welch, D.F., Carroll, K.C., Hofmeister, E.K., Persing, D.H., Robison, D.A., Steigerwalt, A.G., Brenner, D.J., Isolation of a new subspecies, Bartonella vinsonii subsp. Arupensis, from a cattle rancher: Identity with isolates found in conjunction with Borrelia burgdorferi and Babesia microti among naturally infected mice (1999) J Clin Microbiol, 37, pp. 2598-2601Probert, W., Louie, J.K., Tucker, J.R., Longoria, R., Hogue, R., Moler, S., Graves, M., Fritz, C.L., Meningitis due to a "Bartonella washoensis"-like human pathogen (2009) J Clin Microbiol, 47, pp. 2332-2335. , http://dx.doi.org/10.1128/JCM.00511-09Kosoy, M., Morway, C., Sheff, K.W., Bai, Y., Colborn, J., Chalcraft, L., Dowell, S.F., Petersen, L.R., Bartonella tamiae sp. Nov., a newly recognized pathogen isolated from three human patients from Thailand (2008) J Clin Microbiol, 46, pp. 772-775. , http://dx.doi.org/10.1128/JCM.02120-07Maggi, R.G., Kosoy, M., Mintzer, M., Breitschwerdt, E.B., Isolation of Candidatus Bartonella melophagi from human blood (2009) Emerg Infect Dis, 15, pp. 66-68. , http://dx.doi.org/10.3201/eid1501.081080Lin, E.Y., Tsigrelis, C., Baddour, L.M., Lepidi, H., Rolain, J.M., Patel, R., Raoult, D., Candidatus Bartonella mayotimonensis and endocarditis (2010) Emerg Infect Dis, 16, pp. 500-503. , http://dx.doi.org/10.3201/eid1603.081673Breitschwerdt, E.B., Maggi, R.G., Cadenas, M.B., De Paiva Diniz, P.P., A groundhog, a novel Bartonella sequence, and my father's death (2009) Emerg Infect Dis, 15, pp. 2080-2086. , http://dx.doi.org/10.3201/eid1512.AD151

Measuring Electrical And Mechanical Properties Of Red Blood Cells With Double Optical Tweezers

Red blood cell (RBC) aggregation in the blood stream is prevented by the zeta potential created by its negatively charged membrane. There are techniques, however, to decrease the zeta potential and allow cell agglutination, which are the basis of most of antigen-antibody tests used in immunohematology. We propose the use of optical tweezers to measure membrane viscosity, adhesion, zeta potential, and the double layer thickness of charges (DLT) formed around the cell in an electrolytic solution. For the membrane viscosity experiment, we trap a bead attached to RBCs and measure the force to slide one RBC over the other as a function of the velocity. Adhesion is quantified by displacing two RBCs apart until disagglutination. The DLT is measured using the force on the bead attached to a single RBC in response to an applied voltage. The zeta potential is obtained by measuring the terminal velocity after releasing the RBC from the trap at the last applied voltage. We believe that the methodology proposed here can provide information about agglutination, help to improve the tests usually performed in transfusion services, and be applied for zeta potential measurements in other samples. © 2008 Society of Photo-Optical Instrumentation Engineers.131Eylar, E.H., Madoff, M.A., Brody, O.V., Oncley, J.L., The contribution of sialic acid to the surface charge of the erythrocyte (1962) J. Biol. Chem., 237, pp. 1992-2000. , jbc JBCHA3 0021-9258Pollack, W., Reckel, R.P., A reappraisal of the forces involved in hemagglutination (1977) International Archives of Allergy and Applied Immunology, 54 (1), pp. 29-42Stratton, F., Rawlinson, V.I., Gunson, H.H., Phillips, P.K., The role of zeta potential in Rh agglutination (1973) Vox Sang., 24, pp. 273-279. , 7rVOSAAD 0042-9007Luner, S.J., Sturgeon, P., Szklarek, K., McQuiston, D.T., Effects of proteases and neuramidase on RBC surface charge and agglutination (1975) Vox Sang., 28, pp. 184-199. , 7rVOSAAD 0042-9007Zhu, C., Bao, G., Wang, N., Cell Mechanics: Mechanical response, cell adhesion, and molecular deformation (2000) Annu. Rev. Biomed. Eng., 2, pp. 189-226. , bb3 ARBEF7 1523-9829 10.1146/annurev.bioeng.2.1.189Fontes, A., Giorgio, S., De Castro Jr., A.B., Neto, V.M., De Pozzo, L.Y., Marques, G.P., Barbosa, L.C., Cesar, C.L., Determination of Femto Newton forces and fluid viscosity using optical tweezers - Application to Leishmania amazonensis (2005) Progress in Biomedical Optics and Imaging - Proceedings of SPIE, 5699, pp. 419-425. , DOI 10.1117/12.586427, 59, Imaging, Manipulation, and Analysis of Biomolecules and Cells: Fundamentals and Applications IIIChelidze, T., Dielectric spectroscopy of blood (2002) Journal of Non-Crystalline Solids, 305 (1-3), pp. 285-294. , DOI 10.1016/S0022-3093(02)01101-8, PII S0022309302011018Saffman, P.G., Delbruck, M., Brownian motion in biological membranes (1975) Proc. Natl. Acad. Sci. U.S.A., 72, pp. 3111-3113. , pna PNASA6 0027-8424 10.1073/pnas.72.8.3111Dimova, R., Danov, K., Pouligny, B., Ivanov, I.B., Drag of a solid particle trapped in a thin film or at an interface: Influence of surface viscosity and elasticity (2000) Journal of Colloid and Interface Science, 226 (1), pp. 35-43. , DOI 10.1006/jcis.2000.6710Hochmuth, R.M., Worthy, P.R., Evans, E.A., Red cell extensional recovery and the determination of membrane viscosity (1979) Biophysical Journal, 26 (1), pp. 101-114Sze, A., Erickson, D., Ren, L., Li, D., Zeta-potential measurement using the Smoluchowski equation and the slope of the current-time relationship in electroosmotic flow (2003) Journal of Colloid and Interface Science, 261 (2), pp. 402-410. , DOI 10.1016/S0021-9797(03)00142-5Hunter, R.J., (1981) Zeta Potential in Colloid Science, , Academic Press, New YorkPollack, W., Hager, H.J., Reckel, R., Toren, D.A., Singher, H.O., A study of the forces involved in the second stage of hemagglutination (1965) Transfusion (Paris), 5, pp. 158-183. , 7m8 TRNFA2 0372-1248Hymer, W.C., Barlow, G.H., Blaisdell, S.J., Continuous flow electrophoretic separation of proteins and cells from mammalian tissues (1987) Cell Biophys., 10, pp. 61-85. , cbi CBIODE 0163-4992Hashimoto, N., Fujita, S., Yokoyama, T., Ozawa, V., Kingetsu, I., Kurosaka, D., Sabolovic, D., Schuett, W., Cell electrophoretic mobility and glycerol lysis of human erythrocytes in various diseases (1998) Electrophoresis, 19 (7), pp. 1227-1230. , DOI 10.1002/elps.1150190726Carter, H.B., Coffey, D.S., Cell-surface charge in predicting metastatic potential of aspirated cells from the dunning rat prostatic adenocarcinoma model (1988) J. Urol. (Baltimore), 140, pp. 173-175. , jur JOURAA 0022-5347Guck, J., Schinkinger, S., Lincoln, B., Wottawah, F., Ebert, S., Romeyke, M., Lenz, D., Bilby, C., Optical deformability as an inherent cell marker for testing malignant transformation and metastatic competence (2005) Biophysical Journal, 88 (5), pp. 3689-3698. , DOI 10.1529/biophysj.104.04547

Oral Involvement In Chronic Graft Versus Host Disease: A Prospective Study Of 19 Brazilian Patients

Chronic graft versus host disease (cGVHD) is the most common late complication of allogeneic bone marrow transplantation. The oral cavity is the most common site of cGVHD involvement. This study sought to investigate the incidence of oral cGVHD, as well as the disease's impact on a patient's quality of life and the kind of lesions that resulted. Nineteen patients with cGVHD received a medical and dental evaluation; 18 (94.7%) had oral lesions. Nine patients (47.3%) demonstrated xerostomia and 6 (35.2%) demonstrated dysphagia. Six patients (35.2%) had a lichenoid clinical form of cGVHD in the oral cavity, 6 (35.2%) had an atrophic-ulcerative clinical form, 3 (17.6%) had a hyperceratotic clinical form, and 2 (10.5%) had mixed forms. The results demonstrated predominance of lichenoid and ulcerative-atrophic forms with similar incidence of these lesions. No factor that could contribute to the severity of cGVHD oral lesions was found.5514851Storb, R., Thomas, E.D., Allogeneic bone-marrow transplantation (1983) Immunol Rev, 71, pp. 77-102Hunter, A.E., Haynes, A.P., Russell, N.H., Bone marrow transplantation: Current situation, complications and prevention (1995) J Antimicrob Chemother, 36 (SUPPL. B), pp. 119-133Sullivan, K.M., Acute and chronic graft-versus-host disease in man (1986) Int J Cell Cloning, 4 (SUPPL. 1), pp. 42-93Ferrara, J.L., Deeg, H.J., Graft-versus-host disease (1991) N Engl J Med, 324, pp. 667-674Woo SB, Lee SJ, Schubert MM. Graft-vs.-host disease. Crit Rev Oral Biol Med 1997;8:201-216Sedghizadeh, P.P., Alien, C.M., Anderson, K.E., Kim, D.H., Kalmar, J.R., Lang, J.C., Oral graft-versus-host disease and programmed cell death: Pathogenetic and clinical correlates (2004) Oral Surg Oral Med Oral Pathol Oral Radiol Endod, 97, pp. 491-498Couriel, D., Caldera, H., Champlin, R., Komanduri, K., Acute graft-versus-host disease: Pathophysiology, clinical manifestations, and management (2004) Cancer, 101, pp. 1936-1946Sullivan, K.M., Shulman, H.M., Storb, R., Weiden, P.L., Witherspoon, R.P., McDonald, G.B., Schubert, M.M., Thomas, E.D., Chronic graft-versus-host disease in 52 patients: Adverse natural course and successful treatment with combination immunosuppression (1981) Blood, 57, pp. 267-276Eggleston, T.I., Ziccardi, V.B., Lumerman, H., Graft-versus-host disease. Case report and discussion (1998) Oral Surg Oral Med Oral Pathol Oral Radiol Endod, 86, pp. 692-696Schubert, M.M., Sullivan, K.M., Recognition, incidence, and management of oral graft-versus-host disease (1990) NCI Monogr, 9, pp. 135-143Dominguez, R.A., Aznar, M.T., Barberia, L.E., Cabrera, S.E., Oral manifestations of graft versus host disease. Case report (2003) Med Oral, 8, pp. 361-365Nicolatou-Galitis, O., Kitra, V., Van Vliet-Constantinidou, C., Peristeri, J., Goussetis, E., Petropoulos, D., Grafakos, S., The oral manifestations of chronic graft-versus-host disease (cGVHD) in paediatric allogeneic bone marrow transplant recipients (2001) J Oral Pathol Med, 30, pp. 148-153Hiroki, A., Nakamura, S., Shinohara, M., Oka, M., Significance of oral examination in chronic graft-versus-host disease (1994) J Oral Pathol Med, 23, pp. 209-215Rodu, B., Gockerman, J.P., Oral manifestations of the chronic graft-v-host reaction (1983) JAMA, 249, pp. 504-507Nagler, R., Marmary, Y., Krausz, Y., Chisin, R., Markitziu, A., Nagler, A., Major salivary gland dysfunction in human acute and chronic graft-versus-host disease (GVHD) (1996) Bone Marrow Transplant, 17, pp. 219-224LeVeque, F.G., An unusual presentation of chronic graft versus host disease in an unrelated bone marrow transplantation (1990) Oral Surg Oral Med Oral Pathol, 69, pp. 581-584Lee, L., Miller, P.A., Maxymiw, W.G., Messner, H.A., Rotstein, L.E., Intraoral pyogenic granuloma after allogeneic bone marrow transplant. Report of three cases (1994) Oral Surg Oral Med Oral Pathol, 78, pp. 607-610Atkinson, K., Horowitz, M.M., Gale, R.P., Lee, M.B., Rimm, A.A., Bortin, M.M., Consensus among bone marrow transplanters for diagnosis, grading and treatment of chronic graft-versus-host disease. Committee of the International Bone Marrow Transplant Registry (1989) Bone Marrow Transplant, 4, pp. 247-254Franca, C.M., Domingues-Martins, M., Volpe, A., Pallotta Filho, R., Soares de Araujo, N., Severe oral manifestations of chronic graft-vs.-host disease. J Am Dent Assoc (2001), p. 132. , 1124-U27Ratanatharathorn, V., Ayash, L., Lazarus, H.M., Fu, J., Uberti, J.P., Chronic graft-versus-host disease: Clinical manifestation and therapy (2001) Bone Marrow Transplant, 28, pp. 121-129Alborghetti, M.R., Correa, M.E., Adam, R.L., Metze, K., Coracin, F.L., de Souza, C.A., Cintra, M.L., Late effects of chronic graft-vs.-host disease minor salivary glands (2005) J Oral Pathol Med, 34, pp. 486-493Nakamura, S., Hiroki, A., Shinohara, M., Gondo, H., Ohyama, Y., Mouri, T., Sasaki, M., Niho, Y., Oral involvement in chronic graft-versus-host disease after allogeneic bone marrow transplantation (1996) Oral Surg Oral Med Oral Pathol, 82, pp. 556-563Woo, S., Sonis, S.T., Monopoli, M.M., Sonis, A.L., A longitudinal study of oral ulcerative mucositis in bone marrow transplant recipients (1993) Cancer, 72, pp. 1612-1617Demarosi, F., Bez, C., Sardella, A., Lodi, G., Carrassi, A., Oral involvement in chronic graft-vs-host disease following allogenic bone marrow transplantation (2002) Arch Dermatol, 138, pp. 842-84

High Fluorescent And Stable Semiconductor Quantum Dots For Red Blood Cells Labeling

We present a simple and efficient method for marking living human red blood cells using CdS (Cadmium Sulfide) quantum dots (QDs). The nanocrystals were obtained via colloidal synthesis in aqueous medium with final pH=7 using sodium polyphosphate as the stabilizing agent. The methodology implementation is simple, do not requires additional capping layers nor narrow size QDs distribution. The synthesized nanoparticles were conjugated to monoclonal A anti-body. The resulting conjugates QDs/anti-A were incubated with human erythrocytes of blood groups A and O for 30 min at 37°C. The living cells in contact with the quantum dots maintained their properties for several days showing the low level of citotoxicity of the quantum dots. The conjugation of CdS QDs/anti-A show simultaneous red and green fluorescence when excited with 543 and 488 nm respectively. The efficiency of the conjugation QDs/anti-body to the erythrocytes, for each system, was monitored by confocal microscopy. The comparative analysis of the micrographs was done with the luminescence intensity maps of the samples obtained under constant capture conditions, such as, pinhole, filters, beam splitters and photomultiplier gain. The conjugates QDs/anti-A intensely marked group A erythrocytes and did not show any luminescence for group O erythrocytes, showing the sensitivity of the labeling procedure. In conclusion, we show the viability of the use of high luminescent and stable quantum dots as fluorescent labels for human erythrocytes with a methodology of simple implementation and the possibility to use them to distinguish different blood groups.57048692Borchert, H., Talapin, D.V., McGinley, C., High resolution photoemission study of CdSe and CdSe/ZnS core-shell nanocrystals (2003) J. Chem. Phys., 271, pp. 1800-1807Alivisatos, A.P., Semiconductor clusters, nanocrystals, and quantum dots (1996) Science, 271, pp. 933-937Nowak, C., Döllefeld, H., Eychmüller, A., Innershell absorption spectroscopy on CdS: Free clusters and nanocrystals (2001) J. Chem. Phys., 114, pp. 489-494Eychmuller, A., Structure and photophysics of semiconductor nanocrystals (1996) J. Phys Chem. B., 59, pp. 13226-13239Bruchez Jr., M., Moronne, M., Gin, P., Weiss, S., Alivisatos, A.P., Semiconductor nanocrystals as fluorescent biological labels (1998) Science, 281, pp. 2013-2016Chan, W.C.W., Nie, S., Quantum dots for ultrasensitive biological detection (1998) Science, 281, pp. 2016-2018Ballou, B., Lagerholm, B.C., Ernst, L.A., Bruchez, M.P., Waggoner, A.S., Noninvasive imaging of quantum dots in mice (2004) Bioconjug. Chem., 15, pp. 79-86Gao, X., Nie, S., Molecular profiling of single cells and tissue specimens with quantum dots (2003) Trends Biotechnol., 21 (9), pp. 371-373Äkerman, M.E., Chan, W.C.W., Laakkonen, P., Bhatia, S.N., Ruoslahti, E., Nanocrystal targeting in vivo (2002) Appl. Biol. Sci., 99 (20), pp. 12617-12621Chan, W.C.W., Maxwell, D.J., Gao, X., Bailey, R.E., Han, M., Shuming, N., (2002) Curr. Op. Bioteh., 13 (1), pp. 40-46Mattheakis, L.C., Dias, J.M., Choi, Y., Gong, J., Bruchez, M.P., Liu, J., Wang, E., Optical cding of mammalian cells using semiconductor quantum dot (2004) Anal Biochem., 327, pp. 200-208Tokumasu, F., Dvorak, J., Development and application of quantum dots for immunocytochemistry of human erythrocytes (2003) J Microsc., 211, pp. 256-261Hoshino, A., Hanaki, K.-I., Suzuki, K., Yamamoto, K., Applications of T-lymphoma labeled with fluorescent quantum dots to cells tracing markers in mouse body (2004) Biochem. Biophys. Res. Comm., 314, pp. 46-53Petrov, D.V., Santos, B.S., Pereira, G.A.L., Donegá, C.M., Size and band-gap dependences of the hiperpolarizability of CdxZn1-xS nanocrystals (2002) J. Phys. Chem. B, 106 (21), pp. 5325-5334Daniels, G., (2002) Human Blood Groups, , Oxford, UK: Blackwell ScienceTaylor, D.L., Waggoner, A.S., Murphy, R.F., Lanni, F., Birge, R.R., (1986) Applications of Fluorescent in the Biomedical Sciences, , New York, USA: Alan R. Liss, IncLarson, D.R., Zipfel, W.R., Williams, R.M., Clark, S.W., Bruchez, M.P., Wise, F.W., Webb, W.W., Water soluble quantum dots for multiphoton fluorescence imaging in vivo (2003) Science, 300, pp. 1434-143

Red Blood Cell Membrane Viscoelasticity, Agglutination And Zeta Potential Measurements With Double Optical Tweezers

The red blood cell (RBC) viscoelastic membrane contains proteins and glycolproteins embedded in, or attached, to a fluid lipid bilayer and are negatively charged, which creates a repulsive electric (zeta) potential between the cells and prevents their aggregation in the blood stream. There are techniques, however, to decrease the zeta potential to allow cell agglutination which are the basis of most of the tests of antigen-antibody interactions in blood banks. This report shows the use of a double optical tweezers to measure RBC membrane viscosity, agglutination and zeta potential. In our technique one of the optical tweezers trap a silica bead that binds strongly to a RBC at the end of a RBCs rouleaux and, at the same time, acts as a pico-Newton force transducer, after calibration through its displacement from the equilibrium position. The other optical tweezers trap the RBC at the other end. To measure the membrane viscosity the optical force is measured as a function of the velocity between the RBCs. To measure the adhesion the tweezers are slowly displaced apart until the RBCs disagglutination happens. The RBC zeta potential is measured in two complimentary ways, by the force on the silica bead attached to a single RBC in response to an applied electric field, and the conventional way, by the measurement of terminal velocity of the RBC after released from the optical trap. These two measurements provide information about the RBC charges and, also, electrolytic solution properties. We believe this can improve the methods of diagnosis in blood banks.6088Eylar, E.H., Madoff, M.A., Brody, O.V., Oncley, J.L., The contribution of sialic acid to the surface charge of the erythrocyte (1962) J. Biol. Chem., 237, pp. 1992-2000Pollack, W., Reckel, R.P., A reappraisal of the forces involved in Hemagglutination (1977) Int Archs Allergy Appl. Immun., 54, pp. 29-42Ashkin, A., Dziedzic, J.M., Bjorkholm, J.E., Chu, S., Observation of a single-beam gradient force trap for dielectric particles (1986) Opt. Lett., 11, pp. 288-290Ashkin, A., Dziedzic, J.M., Optical trapping and manipulation of viruses and bacteria (1987) Science, 235, pp. 1517-1520Grier, D.G., A revolution in optical manipulation (2003) Nature, 424, pp. 810-816Zhu, C., Bao, G., Wang, N., Cell mechanics: Mechanical response, cell adhesion, and molecular deformation (2000) Annu. Rev. Biomed. Eng., 2, pp. 189-226Neuman, K.C., Block, S., Optical trapping (2004) Rev. Sci. Instrum., 75, pp. 2787-2809Saffman, P.G., Delbruck, M., Brownian motion in biological membranes (1975) Proc. Nat. Acad. Sci. USA, 72, pp. 3111-3113Dimova, R., Danov, K., Pouligny, B., Ivanov, I.B., Drag of a solid particle trapped in a thin film or at an interface: Influence of surface viscosity and elasticity (2000) J. Colloid and Interface Science, 226, pp. 35-43Hochmuth, R., Worthy, P., Evans, E., Red cell extensional recovery and the determination of membrane viscosity (1979) Biophys. J., 26, pp. 101-114Sze, A., Erickson, D., Ren, L., Li, D., Zeta-potential measurement using the Smoluchowski equation and the slope of the current-time relationship in electroosmotic flow (2003) J. Colloid and Interface Science, 261, pp. 402-410Hunter, R.J., (1981) Zeta Potential in Colloid Science, , Academic Press, New YorkPollack, W., Hager, H.J., Reckel, R., Toren, D.A., Singher, H.O., A study of the forces involved in the second stage of hemaggltination (1965) Transfusion, 5, pp. 158-183Chelidze, T., Dielectric spectroscopy of blood (2002) J. Non-crystalline Solids, 305, pp. 285-294Hymer, W.C., Barlow, G.H., Blaisdell, S.J., Continuous flow electrophoretic separation of proteins and cells from mammalian tissues (1987) Cell Biophys., 10, pp. 61-85Hashimoto, N., Fujita, S., Yokoyama, T., Cell electrophoretic mobility and glycerol lysis of human erythrocytes in various diseases (1998) Electrophoresis, 19, pp. 1227-123

Bartonella Spp. Bacteremia In Blood Donors From Campinas, Brazil

Bartonella species are blood-borne, re-emerging organisms, capable of causing prolonged infection with diverse disease manifestations, from asymptomatic bacteremia to chronic debilitating disease and death. This pathogen can survive for over a month in stored blood. However, its prevalence among blood donors is unknown, and screening of blood supplies for this pathogen is not routinely performed. We investigated Bartonella spp. prevalence in 500 blood donors from Campinas, Brazil, based on a cross-sectional design. Blood samples were inoculated into an enrichment liquid growth medium and sub-inoculated onto blood agar. Liquid culture samples and Gram-negative isolates were tested using a genus specific ITS PCR with amplicons sequenced for species identification. Bartonella henselae and Bartonella quintana antibodies were assayed by indirect immunofluorescence. B. henselae was isolated from six donors (1.2%). Sixteen donors (3.2%) were Bartonella-PCR positive after culture in liquid or on solid media, with 15 donors infected with B. henselae and one donor infected with Bartonella clarridgeiae. Antibodies against B. henselae or B. quintana were found in 16% and 32% of 500 blood donors, respectively. Serology was not associated with infection, with only three of 16 Bartonella-infected subjects seropositive for B. henselae or B. quintana. Bartonella DNA was present in the bloodstream of approximately one out of 30 donors from a major blood bank in South America. Negative serology does not rule out Bartonella spp. infection in healthy subjects. Using a combination of liquid and solid cultures, PCR, and DNA sequencing, this study documents for the first time that Bartonella spp. bacteremia occurs in asymptomatic blood donors. Our findings support further evaluation of Bartonella spp. transmission which can occur through blood transfusions.91Breitschwerdt, E.B., Maggi, R.G., Chomel, B.B., Lappin, M.R., Bartonellosis: an emerging infectious disease of zoonotic importance to animals and human beings (2010) J Vet Emerg Crit Care, 20, pp. 8-30Chomel, B.B., Boulouis, H.J., Breitschwerdt, E.B., Kasten, R.W., Vayssier-Taussat, M., Ecological fitness and strategies of adaptation of Bartonella species to their hosts and vectors (2009) Vet Res, 40, p. 29Boulouis, H.J., Chang, C.C., Henn, J.B., Kasten, R.W., Chomel, B.B., Factors associated with the rapid emergence of zoonotic Bartonella infections (2005) Vet Res, 36, pp. 383-410Maggi, R.G., Mozayeni, B.R., Pultorak, E.L., Hegarty, B.C., Bradley, J.M., Bartonella spp. bacteremia and rheumatic symptoms in patients from Lyme disease-endemic region (2012) Emerg Infect Dis, 18, pp. 783-791Breitschwerdt, E.B., Maggi, R.G., Lantos, P.M., Woods, C.W., Hegarty, B.C., Bartonella vinsonii subsp. berkhoffii and Bartonella henselae bacteremia in a father and daughter with neurological disease (2010) Parasit Vectors, 3, p. 29Garcia-Caceres, U., Garcia, F.U., Bartonellosis. An immunodepressive disease and the life of Daniel Alcides Carrion (1991) Am J Clin Pathol, 95, pp. S58-66Pitassi, L.H., Cintra, M.L., Ferreira, M.R., Magalhaes, R.F., Velho, P.E., Blood cell findings resembling Bartonella spp (2010) Ultrastruct Pathol, 34, pp. 2-6Maggi, R.G., Mascarelli, P.E., Pultorak, E.L., Hegarty, B.C., Bradley, J.M., Bartonella spp. bacteremia in high-risk immunocompetent patients (2011) Diagn Microbiol Infect Dis, 71, pp. 430-437Harms, A., Dehio, C., Intruders below the radar: molecular pathogenesis of Bartonella spp (2012) Clin Microbiol Rev, 25, pp. 42-78Pulliainen, A.T., Dehio, C., Persistence of Bartonella spp. stealth pathogens: from subclinical infections to vasoproliferative tumor formation (2012) FEMS Microbiol Rev, 36, pp. 563-599Duncan, A.W., Maggi, R.G., Breitschwerdt, E.B., A combined approach for the enhanced detection and isolation of Bartonella species in dog blood samples: pre-enrichment liquid culture followed by PCR and subculture onto agar plates (2007) J Microbiol Methods, 69, pp. 273-281Magalhaes, R.F., Pitassi, L.H., Salvadego, M., de Moraes, A.M., Barjas-Castro, M.L., Bartonella henselae survives after the storage period of red blood cell units: is it transmissible by transfusion? (2008) Transfus Med, 18, pp. 287-291Pitassi, L.H., Magalhaes, R.F., Barjas-Castro, M.L., de Paula, E.V., Ferreira, M.R., Bartonella henselae infects human erythrocytes (2007) Ultrastruct Pathol, 31, pp. 369-372Kordick, D.L., Brown, T.T., Shin, K., Breitschwerdt, E.B., Clinical and pathologic evaluation of chronic Bartonella henselae or Bartonella clarridgeiae infection in cats (1999) J Clin Microbiol, 37, pp. 1536-1547Oliveira, A.M., Maggi, R.G., Woods, C.W., Breitschwerdt, E.B., Suspected needle stick transmission of Bartonella vinsonii subspecies berkhoffii to a veterinarian (2010) J Vet Intern Med, 24, pp. 1229-1232Lin, J.W., Chen, C.M., Chang, C.C., Unknown fever and back pain caused by Bartonella henselae in a veterinarian after a needle puncture: a case report and literature review (2011) Vector Borne Zoonotic Dis, 11, pp. 589-591Arvand, M., Schad, S.G., Isolation of Bartonella henselae DNA from the peripheral blood of a patient with cat scratch disease up to 4 months after the cat scratch injury (2006) J Clin Microbiol, 44, pp. 2288-2290Lantos, P.M., Maggi, R.G., Ferguson, B., Varkey, J., Park, L.P., Detection of Bartonella species in the blood of veterinarians and veterinary technicians: a newly recognized occupational hazard? 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