Expression of a hantavirus N protein and its efficacy as antigen in immune assays

Hantavirus cardiopulmonary syndrome (HCPS) has been recognized as an important public heath problem. Five hantaviruses associated with HCPS are currently known in Brazil: Juquitiba, Araraquara, Laguna Negra-like, Castelo dos Sonhos, and Anajatuba viruses. The laboratory diagnosis of HCPS is routinely carried out by the detection of anti-hantavirus IgM and/or IgG antibodies. The present study describes the expression of the N protein of a hantavirus detected in the blood sample of an HCPS patient. The entire S segment of the virus was amplified and found to be 1858 nucleotides long, with an open reading frame of 1287 nucleotides that encodes a protein of 429 amino acids. The nucleotide sequence described here showed a high identity with the N protein gene of Araraquara virus. The entire N protein was expressed using the vector pET200D and the Escherichia coli BL21 strain. The expression of the recombinant protein was confirmed by the detection of a 52-kDa protein by Western blot using a pool of human sera obtained from HCPS patients, and by specific IgG detection in five serum samples of HCPS patients tested by ELISA. These results suggest that the recombinant N protein could be used as an antigen for the serological screening of hantavirus infection.

Effects of systemic administration of ciliary neurotrophic factor on Bax and Bcl-2 proteins in the lumbar spinal cord of neonatal rats after sciatic nerve transection

Ciliary neurotrophic factor (CNTF) is a cytokine that plays a neuroprotective role in relation to axotomized motoneurons. We determined the effect of daily subcutaneous doses of CNTF (1.2 µg/g for 5 days; N = 13) or PBS (N = 13) on the levels of mRNA for Bcl-2 and Bax, as well as the expression and inter-association of Bcl-2 and Bax proteins, and the survival of motoneurons in the spinal cord lumbar enlargement of 2-day-old Wistar rats after sciatic nerve transection. Five days after transection, the effects were evaluated on histological and molecular levels using Nissl staining, immunoprecipitation, Western blot analysis, and reverse transcriptase-polymerase chain reaction. The motoneuron survival ratio, defined as the ratio between the number of motoneurons counted on the lesioned side vs those on the unlesioned side, was calculated. This ratio was 0.77 ± 0.02 for CNTF-treated rats vs 0.53 ± 0.02 for the PBS-treated controls (P < 0.001). Treatment with CNTF modified the level of mRNA, with the expression of Bax RNA decreasing 18% (with a consequent decrease in the level of Bax protein), while the expression of Bcl-2 RNA was increased 87%, although the level of Bcl-2 protein was unchanged. The amount of Bcl-2/Bax heterodimer increased 91% over that found in the PBS-treated controls. These data show, for the first time, that the neuroprotective effect of CNTF on neonatal rat axotomized motoneurons is associated with a reduction in free Bax, due to the inhibition of Bax expression, as well as increased Bcl-2/Bax heterodimerization. Thus, the neuroprotective action of the CNTF on axotomized motoneurons can be related to the inhibition of this apoptotic pathway.

Follicular helper T cell in immunity and autoimmunity

The traditional concept that effector T helper (Th) responses are mediated by Th1/Th2 cell subtypes has been broadened by the recent demonstration of two new effector T helper cells, the IL-17 producing cells (Th17) and the follicular helper T cells (Tfh). These new subsets have many features in common, such as the ability to produce IL-21 and to express the IL-23 receptor (IL23R), the inducible co-stimulatory molecule ICOS, and the transcription factor c-Maf, all of them essential for expansion and establishment of the final pool of both subsets. Tfh cells differ from Th17 by their ability to home to B cell areas in secondary lymphoid tissue through interactions mediated by the chemokine receptor CXCR5 and its ligand CXCL13. These CXCR5+ CD4+ T cells are considered an effector T cell type specialized in B cell help, with a transcriptional profile distinct from Th1 and Th2 cells. The role of Tfh cells and its primary product, IL-21, on B-cell activation and differentiation is essential for humoral immunity against infectious agents. However, when deregulated, Tfh cells could represent an important mechanism contributing to exacerbated humoral response and autoantibody production in autoimmune diseases. This review highlights the importance of Tfh cells by focusing on their biology and differentiation processes in the context of normal immune response to infectious microorganisms and their role in the pathogenesis of autoimmune diseases.

PRODUÇÃO DE IMUNORREAGENTES PARA USO EM UM TESTE IMUNOENZIMÁTICO DE DETECÇÃO DE SALMONELLA EM ALIMENTOS

O objetivo deste trabalho foi o desenvolvimento de imunorreagentes policlonais convenientes para uso em um teste imunoenzimático para detecção rápida de Salmonella em alimentos. Foram produzidos seis anti-soros policlonais anti-Salmonella contendo as aglutininas e, h; 1,6; i; 1,2; f,g,s e m,t. Como antígenos, foram empregadas culturas formolizadas de quatro sorotipos de Salmonella (S. Typhimurium fases 1 e 2, S. Anatum fases 1 e 2, S. Agona monofásica e S. Oranienburg monofásica) cultivadas em caldo tripticase de soja. O teste imunoenzimático utilizado foi o indireto, empregando-se anti-IgG-peroxidase como conjugado e OPD-H2O2 como sistema cromógeno. Os testes imunoenzimáticos foram conduzidos com os anti-soros policlonais individuais absorvidos e não-absorvidos, bem como empregando-se um anti-soro polivalente, correspondente a um "pool" de anti-soros absorvidos e não-absorvidos com culturas puras de Salmonella e outras enterobactérias. A absorção dos soros eliminou as reações cruzadas com todas as enterobactérias testadas nesse estudo, apresentadas tanto por alguns anti-soros individuais quanto pelo polivalente. Entre os seis anti-soros estudados, os que apresentaram melhor desempenho foram o f,g,s não-absorvido e o polivalente absorvido.

Free and total GMP (glycomacropeptide) contents of milk during bovine lactation

Individual milk samples taken every two weeks from parturition to the end of lactation from 34 animals of three different herds and breeds were analyzed for free-GMP. A milk pool of each herd was analyzed for free and total GMP (released from k-casein by the action of rennin) and the data were correlated with sanitary conditions of animal and udder, phase of lactation and milk production. Most udder problems were concentrated near parturition, with few and spaced occurrences of clinical mastitis. The Californian Mastitis Test (CMT) results showed oscillations compatible with the phases of lactation period and environmental conditions. The widest variations in free-GMP occurred as a function of lactation period and as a consequence of clinical or subclinical mastitis. Higher levels were observed at the beginning of lactation (5.87mg L-1 of sialic acid), becoming normal with mean values of about 3.30mg L-1 at the end of the second month, and increasing again during the final third of lactation. On average, the same trends were observed for total GMP released by commercial rennet, beginning with slightly high values (35.59mg L-1), becoming normal by the sixth month with values close to 27.15mg L-1, and rising gradually up to the end of lactation, with 58.35mg L-1 of sialic acid. These results prove to be useful for the correct interpretation of tests applied to milk selection with respect to proteolytic status or even to restrain frauds by the addition of whey to milk.

Weldability of powder bed fusion fabricated stainless steel 316L sheets to cold rolled sheet metal

Weldability of powder bed fusion (PBF) fabricated components has come to discussion in past two years due to resent developments in the PBF technology and limited size of the machines used in the fabrication process. This study concentrated on effects of energy input of welding on mechanical properties and microstructural features of welds between PBF fabricated stainless steel 316L sheets and cold rolled sheet metal of same composition by the means of destructive testing and microscopic analysis. Optical fiber diameter, laser power and welding speed were varied during the experiments that were executed following one variable at a time (OVAT) method. One of the problems of welded PBF fabricated components has been lower elongations at break comparing to conventionally manufactured components. Decreasing energy input of the laser keyhole welding decreased elongations at break of the welded specimens. Ultimate tensile strengths were not affected significantly by the energy input of the welding, but fracturing of the specimens welded using high energy input occurred from the weld metal. Fracturing of the lower energy input welds occurred from the PBF fabricated base metal. Energy input was found to be critical factor for mechanical properties of the welds. Multioriented grain growth and formation of neck at fusion zone boundary on the cold rolled side of the weld was detected and suspected to be result from weld pool flows caused by differences in molten weld pool behaviour between the PBF fabricated and cold rolled sides of the welds.

Effect of pectinolitic enzymes on the physical properties of caja-manga (Spondias cytherea Sonn.) pulp

Cajá-manga, also known as golden apple and hog-plum, is an exotic fruit native from Îles de la Société (French Polynesia), which was first introduced in Brazil in 1985. The pulp of ripe fruit was treated with the commercial enzymatic pool and its effect was evaluated in terms of yield, as well as the physical properties viscosity, turbidity and color (L* values). Response surface methodology was used and three levels were adopted for the independent variables temperature (30, 40, and 50 ºC), incubation time (30, 60 and 90 minutes) and enzyme concentration (0.01, 0.05, 0.09 v/v%). A central composite statistical design was used to guide the experimental work. The enzyme treatment highly increased both juice yield (up to 56%) and color (up to 8.6%) and strongly decreased viscosity (up to 57.4%), clarity (up to 77%) and turbidity (up to 85.5%). Incubation time was the most interacting facto, whereas temperature was the least one. Optimization analysis was carried out to reduce enzyme concentration to a minimum by superposing the contour plots of the tested properties, and the recommended ranges of the variables enzyme concentration, process temperature and incubation time were, respectively, 0.042-0.068%, 47.0-49.0 ºC and 82-90 minutes.

Ocurrence of Vibrio spp., positive coagulase staphylococci and enteric bacteria in oysters (Crassostrea gigas) harvested in the south bay of Santa Catarina island, Brazil

The aim of this study was to assess the contamination of oysters (Crassostrea gigas), harvested in six different regions of the South Bay of Santa Catarina Island, with Coliforms at 45 ºC, Escherichia coli, Vibrio spp., positive coagulase staphylococci, and Salmonella sp. over a period of one year. One hundred eighty oyster samples were collected directly from their culture sites and analyzed. Each sample consisted of a pool of 12 oysters. All of the samples analyzed showed absence of Salmonella, 18 (10%) samples showed presence of Escherichia coli, 15 (8.3%) samples were positive for V. alginolyticus, and Vibriocholerae was detected in 4 samples (2.2%). The counts of positive-coagulase staphylococci varied from <10 to 1.9 x 102 CFU.g-1, whereas the counts of Coliforms at 45 ºC and E. coli ranged from <3 to 1.5 x 102 MPN.g-1 and <3 and 4.3 x 10 MPN.g-1, respectively. Counts of V. parahaemolyticus and V. vulnificus ranged between <3 and 7 MPN.g-1, for both microorganisms. This suggests the need for monitoring these Vibrios contamination in oysters. Based on the results of the microbiological assays, the samples analyzed showed acceptable bacteriological quality, i.e., they were within the parameters established by Brazilian Legislation.

Coolability of porous core debris beds : Effects of bed geometry and multi-dimensional flooding

This thesis addresses the coolability of porous debris beds in the context of severe accident management of nuclear power reactors. In a hypothetical severe accident at a Nordic-type boiling water reactor, the lower drywell of the containment is flooded, for the purpose of cooling the core melt discharged from the reactor pressure vessel in a water pool. The melt is fragmented and solidified in the pool, ultimately forming a porous debris bed that generates decay heat. The properties of the bed determine the limiting value for the heat flux that can be removed from the debris to the surrounding water without the risk of re-melting. The coolability of porous debris beds has been investigated experimentally by measuring the dryout power in electrically heated test beds that have different geometries. The geometries represent the debris bed shapes that may form in an accident scenario. The focus is especially on heap-like, realistic geometries which facilitate the multi-dimensional infiltration (flooding) of coolant into the bed. Spherical and irregular particles have been used to simulate the debris. The experiments have been modeled using 2D and 3D simulation codes applicable to fluid flow and heat transfer in porous media. Based on the experimental and simulation results, an interpretation of the dryout behavior in complex debris bed geometries is presented, and the validity of the codes and models for dryout predictions is evaluated. According to the experimental and simulation results, the coolability of the debris bed depends on both the flooding mode and the height of the bed. In the experiments, it was found that multi-dimensional flooding increases the dryout heat flux and coolability in a heap-shaped debris bed by 47–58% compared to the dryout heat flux of a classical, top-flooded bed of the same height. However, heap-like beds are higher than flat, top-flooded beds, which results in the formation of larger steam flux at the top of the bed. This counteracts the effect of the multi-dimensional flooding. Based on the measured dryout heat fluxes, the maximum height of a heap-like bed can only be about 1.5 times the height of a top-flooded, cylindrical bed in order to preserve the direct benefit from the multi-dimensional flooding. In addition, studies were conducted to evaluate the hydrodynamically representative effective particle diameter, which is applied in simulation models to describe debris beds that consist of irregular particles with considerable size variation. The results suggest that the effective diameter is small, closest to the mean diameter based on the number or length of particles.

Family rivalry in the testis: Retinoblastoma protein and E2F transcription factors in testicular development and adult spermatogenesis

Disorders of male reproductive health are becoming increasingly prevalent globally. These defects, ranging from decreasing sperm counts to an increasing rate of infertility and testicular cancer, have a common origin in the early phases of testicular development, but the exact mechanisms that cause them remain unknown. Testicular development and adult spermatogenesis are complex processes in which different cell types undergo mitosis, meiosis, differentiation and apoptosis. The retinoblastoma protein family and its associated E2F transcription factors are key regulators of these cellular events. In the present study, the functions of these factors in postnatal testicular development and adult spermatogenesis were explored using different animal models. In addition, a new application of flow cytometry to study testicular cell dynamics was developed. An ablation of retinoblastoma protein in mouse Sertoli cells resulted in their cell cycle re-entry in adult testes, dedifferentiation and a severe spermatogenic defect. We showed that deregulated E2F3 contributed to these changes. Our results indicated that the E2F1 transcription factor is critical for the control of apoptosis in the developing postnatal testis. In the adult testis, E2F1 controls the maintenance of the spermatogonial stem cell pool, in addition to inhibiting apoptosis of spermatocytes. In summary, this study elucidated the complex interdependencies of the RB and E2F transcription factor families in the control of postnatal testicular development and adult spermatogenesis. Furthermore, this study provided a new methodology for the analysis of testicular cells.

Mineraalien rikastusprosessin vesijalanjälki

Mineraalien rikastamiseen käytetään useita fysikaalisia ja kemiallisia menetelmiä. Prosessi sisältää malmin hienonnuksen, rikastuksen ja lopuksi vedenpoistamisen rikastelietteestä. Malmin rikastamiseen käytetään muun muassa vaahdotusta, liuotusta, magneettista rikastusta ja tiheyseroihin perustuvia rikastusmenetelmiä. Rikastuslietteestä voidaan poistaa vettä sakeuttamalla ja suodattamalla. Rikastusprosessin ympäristövaikutuksia voidaan arvioida laskemalla tuotteen vesijalanjälki, joka kertoo valmistamiseen kulutetun veden määrän. Tässä kirjallisuustyössä esiteltiin mineraalien käsittelymenetelmiä sekä prosessijätevesien puhdistusmenetelmiä. Kirjallisuuslähteiden pohjalta selvitettiin Pyhäsalmen kaivoksella valmistetun kuparianodin vesijalanjälki sekä esitettiin menetelmiä, joilla prosessiin tarvittavan raakaveden kulutusta voitaisiin vähentää. Pyhäsalmella kuparirikasteesta valmistetun kuparianodin vesijalanjälki on 240 litraa H2O ekvivalenttia tuotettua tonnia kohden. Pyhäsalmen prosessin raakaveden kulutusta voidaan vähentää lisäämällä sisäistä vedenkierrätystä. Kalsiumsulfaatin saostuminen putkiin ja pumppuihin on ilmentynyt ongelmaksi vedenkierrätyksen lisäämisessä. Kalsiumsulfaattia voidaan erottaa vedestä membraaneihin, ioninvaihtoon ja sähkökemiaan perustuvilla tekniikoilla. Vaihtoehdossa, jossa johdetaan kaikista kolmesta vaahdotuksesta saatavat rikastuslietteen ja rikastushiekan sakeutuksien ylitteet sekä suodatuksien suodosvedet samaan vedenkäsittelyyn voidaan kattaa arviolta noin 65 % koko veden tarpeesta. Raakavettä säästetään vuodessa 3,4 Mm^3 ja samalla rikastushiekka-altaiden tarvittava koko pienenee, joka vähentää ympäristöriskejä.

Alternative electron transfer routes involved in photoprotection of cyanobacteria

In oxygenic photosynthesis, the highly oxidizing reactions of water splitting produce reactive oxygen species (ROS) and other radicals that could damage the photosynthetic apparatus and affect cell viability. Under particular environmental conditions, more electrons are produced in water oxidation than can be harmlessly used by photochemical processes for the reduction of metabolic electron sinks. In these circumstances, the excess of electrons can be delivered, for instance, to O2, resulting in the production of ROS. To prevent detrimental reactions, a diversified assortment of photoprotection mechanisms has evolved in oxygenic photosynthetic organisms. In this thesis, I focus on the role of alternative electron transfer routes in photoprotection of the cyanobacterium Synechocystis sp. PCC 6803. Firstly, I discovered a novel subunit of the NDH-1 complex, NdhS, which is necessary for cyclic electron transfer around Photosystem I, and provides tolerance to high light intensities. Cyclic electron transfer is important in modulating the ATP/NADPH ratio under stressful environmental conditions. The NdhS subunit is conserved in many oxygenic phototrophs, such as cyanobacteria and higher plants. NdhS has been shown to link linear electron transfer to cyclic electron transfer by forming a bridge for electrons accumulating in the Ferredoxin pool to reach the NDH-1 complexes. Secondly, I thoroughly investigated the role of the entire flv4-2 operon in the photoprotection of Photosystem II under air level CO2 conditions and varying light intensities. The operon encodes three proteins: two flavodiiron proteins Flv2 and Flv4 and a small Sll0218 protein. Flv2 and Flv4 are involved in a novel electron transport pathway diverting electrons from the QB pocket of Photosystem II to electron acceptors, which still remain unknown. In my work, it is shown that the flv4-2 operon-encoded proteins safeguard Photosystem II activity by sequestering electrons and maintaining the oxidized state of the PQ pool. Further, Flv2/Flv4 was shown to boost Photosystem II activity by accelerating forward electron flow, triggered by an increased redox potential of QB. The Sll0218 protein was shown to be differentially regulated as compared to Flv2 and Flv4. Sll0218 appeared to be essential for Photosystem II accumulation and was assigned a stabilizing role for Photosystem II assembly/repair. It was also shown to be responsible for optimized light-harvesting. Thus, Sll0218 and Flv2/Flv4 cooperate to protect and enhance Photosystem II activity. Sll0218 ensures an increased number of active Photosystem II centers that efficiently capture light energy from antennae, whilst the Flv2/Flv4 heterodimer provides a higher electron sink availability, in turn, promoting a safer and enhanced activity of Photosystem II. This intertwined function was shown to result in lowered singlet oxygen production. The flv4-2 operon-encoded photoprotective mechanism disperses excess excitation pressure in a complimentary manner with the Orange Carotenoid Protein-mediated non-photochemical quenching. Bioinformatics analyses provided evidence for the loss of the flv4-2 operon in the genomes of cyanobacteria that have developed a stress inducible D1 form. However, the occurrence of various mechanisms, which dissipate excitation pressure at the acceptor side of Photosystem II was revealed in evolutionarily distant clades of organisms, i.e. cyanobacteria, algae and plants.

Käyttäjien manipulointi internetin yhteisöpalveluissa: Systemaattinen kirjallisuuskatsaus

Internetin yhteisöpalveluiden käyttäjien avoimuus ja sosiaalisuus altistavat heidät monenlaisille riskeille. “Social engineering” eli käyttäjien manipulointi on uhka, joka liittyy informaation hankkimiseen perinteisen kanssakäymisen kautta, mutta yhä enenevissä määrin myös internetissä. Kun kanssakäyminen tapahtuu internetin välityksellä, käyttäjien manipuloijat hyödyntävät yhteisöpalveluita yhteydenpitoon uhrien kanssa sekä paljon käyttäjäinformaatiota sisältävänä alustana. Tämän tutkielman tarkoitus on löytää internetin yhteisöpalveluiden ja käyttäjien manipuloinnin välinen yhteys. Tämä päämäärä saavutettiin etsimällä vastauksia kysymyksiin kuten: Mitkä ovat tyypilliset hyökkäystyypit? Miksi informaatiolla on niin suuri rooli? Mitä seurauksia ilmiöllä on ja miten hyökkäyksiltä on mahdollista suojautua? Vastaukset kysymyksiin löydettiin toteuttamalla systemaattinen kirjallisuuskatsaus. Katsaus muodostui yhdistämällä tärkeimmät löydökset 60 tarkoin valitusta ilmiötä käsittelevästä artikkelista. Käyttäjien manipuloinnin huomattiin olevan hyvin laaja ja monimutkainen ilmiö internetin yhteisöpalveluissa. Huomattiin, että manipulointia ilmenee sivustoilla useissa erilaisissa muodoissa, joita ovat muun muassa tietojen kalastelu, profiilien yhdistäminen, sosiaaliset sovellukset, roskaposti, haitalliset linkit, identiteettivarkaudet, tietovuodot ja erilaiset huijaukset, jotka hyödyntävät sekä ihmisluonnon että sivustojen perusominaisuuksia. Haavoittuvuus ja luottamus havaittiin myös tärkeiksi aspekteiksi, sillä ne yhdistävät informaation merkityksen ja ihmisluonnon, jotka molemmat ovat avaintekijöitä sekä manipuloinnissa että yhteisöpalvelusivustoilla. Vaikka ilmiön seurausten huomattiin olevan negatiivisia niin käyttäjien olemukselle internetissä kuin todellisessakin elämässä, havaittiin myös, että ilmiön ymmärtäminen ja tunnistaminen helpottaa siltä suojautumista

Monitoring Directed Energy Deposition Processes in Additive Manufacturing

The purpose of this study is to find out how laser based Directed Energy Deposition processes can benefit from different types of monitoring. DED is a type of additive manufacturing process, where parts are manufactured in layers by using metallic powder or metallic wire. DED processes can be used to manufacture parts that are not possible to manufacture with conventional manufacturing processes, when adding new geometries to existing parts or when wanting to minimize the scrap material that would result from machining the part. The aim of this study is to find out why laser based DED-processes are monitored, how they are monitored and what devices are used for monitoring. This study has been done in the form of a literature review. During the manufacturing process, the DED-process is highly sensitive to different disturbances such as fluctuations in laser absorption, powder feed rate, temperature, humidity or the reflectivity of the melt pool. These fluctuations can cause fluctuations in the size of the melt pool or its temperature. The variations in the size of the melt pool have an effect on the thickness of individual layers, which have a direct impact on the final surface quality and dimensional accuracy of the parts. By collecting data from these fluctuations and adjusting the laser power in real-time, the size of the melt pool and its temperature can be kept within a specified range that leads to significant improvements in the manufacturing quality. The main areas of monitoring can be divided into the monitoring of the powder feed rate, the temperature of the melt pool, the height of the melt pool and the geometry of the melt pool. Monitoring the powder feed rate is important when depositing different material compositions. Monitoring the temperature of the melt pool can give information about the microstructure and mechanical properties of the part. Monitoring the height and the geometry of the melt pool is an important factor in achieving the desired dimensional accuracy of the part. By combining multiple different monitoring devices, the amount of fluctuations that can be controlled will be increased. In addition, by combining additive manufacturing with machining, the benefits of both processes could be utilized.

NEGOTIATING IN FINNISH GROCERY RETAIL - Negotiation Narratives of S-group, Kesko and Lidl

In this research retail negotiations are explored through the question: What characteristics are distinctive to negotiating in Finnish grocery retail trade? To shed light on the research question I interviewed experienced retail negotiators and mapped out the most important characteristics of the retail negotiations. I described through examples the most prominent challenges negotiators face in their negotiations and elaborated what kind of tools the experienced negotiators use to overcome those challenges. The research results add up to a groundwork frame for retail negotiations with which further research can be more easily directed to any area of interest in the Finnish grocery retail negotiations. The framework can give ideas or frames for further research, or function as a general guideline of factors to consider when negotiating in Finnish retail field. The results were divided into 3 sections: Characteristics, Challenges and Tools. Different negotiation models help negotiators and researchers understand negotiation dynamics. This research adds to that pool by focusing on elements essential to consider specifically in the context of Finnish retail. Finland offered an exceptionally interesting setting to study negotiation, as grocery retail trade in Finland is highly centralized. Especially for those interested understanding a centralized setting such as Finland’s retail field, the framework presented in this research might provide a valuable spectrum of essential negotiation elements. Learning is a lifelong process, but that path can be evened by tuning in on what others have learned during their own endeavors in similar situations. Seasoned negotiators have many stories to tell about negotiating that can be drawn upon and by doing so, we can avoid having to spend time learning the same insights twice. This research drew on narrative, case-research and interviewing to find out how seasoned negotiators in the field of Finnish retail experienced negotiation, what challenges negotiations pose and what tools can be used to overcome them