Plasmablastic myeloma transformation of light-chain multiple myeloma.

Plasmablastic myeloma (PBM) is an uncommon and aggressive morphologic variant of multiple myeloma (MM). The neoplastic immature cells exhibit diverse morphology, posing a diagnostic challenge. The diagnostic criteria for PBM include the identification of ≥ 2% plasmablasts in the bone marrow aspirate. This case describes the incidental finding of a light-chain multiple myeloma (LCMM) transformed into PBM, a phenomenon not previously reported.

Elevated vitamin B12 levels in autoimmune lymphoproliferative syndrome attributable to elevated haptocorrin in lymphocytes.

OBJECTIVE: Identify the etiology of elevated B(12) in autoimmune lymphoproliferative syndrome (ALPS). DESIGN: Peripheral blood of ALPS patients with elevated B(12) and controls were evaluated. RESULTS: Total and holo-haptocorrin (HC) levels were 26- and 23-fold higher in ALPS patients, respectively. No abnormal B(12)-binding proteins were found. Western blot revealed HC in lymphocyte lysates only from ALPS patients. CONCLUSION: Elevated concentrations of B(12) found in ALPS patients were due to increased lymphocyte expression of HC.

Taking a new biomarker into routine use--a perspective from the routine clinical biochemistry laboratory.

There is increasing pressure to provide cost-effective healthcare based on "best practice." Consequently, new biomarkers are only likely to be introduced into routine clinical biochemistry departments if they are supported by a strong evidence base and if the results will improve patient management and outcome. This requires convincing evidence of the benefits of introducing the new test, ideally reflected in fewer hospital admissions, fewer additional investigations and/or fewer clinic visits. Carefully designed audit and cost-benefit studies in relevant patient groups must demonstrate that introducing the biomarker delivers an improved and more effective clinical pathway. From the laboratory perspective, pre-analytical requirements must be thoroughly investigated at an early stage. Good stability of the biomarker in relevant physiological matrices is essential to avoid the need for special processing. Absence of specific timing requirements for sampling and knowledge of the effect of medications that might be used to treat the patients in whom the biomarker will be measured is also highly desirable. Analytically, automation is essential in modern high-throughput clinical laboratories. Assays must therefore be robust, fulfilling standard requirements for linearity on dilution, precision and reproducibility, both within- and between-run. Provision of measurements by a limited number of specialized reference laboratories may be most appropriate, especially when a new biomarker is first introduced into routine practice.

Questões atuais relativas à dosagem e à descrição da excreção urinária de albumina / Current issues in measurement and report of urinary albumin excretion

ANTECEDENTES: A excreção urinária de albumina indica lesão nos rins e é reconhecida como fator de risco para a progressão das doenças renal e cardiovascular. A dosagem da albumina urinária chama a atenção sobre a necessidade clínica de relatos de resultados precisos e claramente descritos. O National Kidney Disease Education Program e a Federação Internacional de Química Clínica e Medicina Laboratorial (IFCC) reuniram-se para avaliar o estado atual das questões pré-analíticas, analíticas e pós-analíticas que afetam as dosagens da albumina na urina e para identificar as áreas que necessitam de melhorias. CONTEÚDO: A química da albumina na urina não é completamente compreendida. Diretrizes atuais recomendam a utilização da relação albumina/creatinina (RAC) como substituta para a coleta de amostras cronometradas de urina, frequentemente inadequadas. Os resultados da RAC são afetados pela preparação do paciente, pela hora do dia da coleta das amostras e não é padronizada. Foram relatadas consideráveis diferenças intermétodos para a dosagem tanto de albumina quanto de creatinina, mas a verdade é desconhecida, porque não existem procedimentos de referência para a dosagem de albumina e não há materiais de referência para qualquer um desses analitos na urina. Os intervalos de referência recomendados para a RAC não consideram as grandes diferenças intergrupos na excreção da creatinina (por exemplo, relacionadas com diferenças em idade, sexo e etnia), nem o aumento contínuo no risco relacionado com a excreção de albumina. DISCUSSÃO: Necessidades clínicas foram identificadas para a padronização de (a) métodos de coleta da urina, (b) dosagens de albumina e de creatinina na urina com base em um sistema de referência completo, (c) relatórios dos resultados dos testes e (d) intervalos de referência para a RAC.

BACKGROUND: Urinary excretion of albumin indicates kidney damage and is recognized as a risk factor for progression of kidney disease and cardiovascular disease. The role of urinary albumin measurements has focused attention on the clinical need for accurate and clearly reported results. The National Kidney Disease Education Program and the IFCC convened a conference to assess the current state of preanalytical, analytical, and postanalytical issues affecting urine albumin measurements and to identify areas needing improvement. CONTENT: The chemistry of albumin in urine is incompletely understood. Current guidelines recommend the use of the albumin/creatinine ratio (ACR) as a surrogate for the error-prone collection of timed urine samples. Although ACR results are affected by patient preparation and time of day of sample collection, neither is standardized. Considerable intermethod differences have been reported for both albumin and creatinine measurement, but trueness is unknown because there are no reference measurement procedures for albumin and no reference materials for either analyte in urine. The recommended reference intervals for the ACR do not take into account the large intergroup differences in creatinine excretion (e.g., related to differences in age, sex, and ethnicity) nor the continuous increase in risk related to albumin excretion. DISCUSSION: Clinical needs have been identified for standardization of (a) urine collection methods, (b) urine albumin and creatinine measurements based on a complete reference system, (c) reporting of test results, and (d) reference intervals for the ACR.

Introduction to urinalysis: historical perspectives and clinical application.

Urinalysis was the first laboratory test performed in medicine and has been used for several thousand years. Today urinalysis continues to be a powerful tool in obtaining crucial information for diagnostic purposes in medicine. Urine is an unstable fluid, and changes to its composition begin to take place as soon as it is voided. As such, collection, storage, and handling are important issues in maintaining the integrity of this specimen. In the laboratory, urine can be characterized by physical appearance, chemical composition, and microscopically. Physical examination of urine includes description of color, odor, clarity, volume, and specific gravity. Chemical examination of urine includes the identification of protein, blood cells, glucose, pH, bilirubin, urobilinogen, ketone bodies, nitrites, and leukocyte esterase. Finally, microscopic examination entails the detection of crystals, cells, casts, and microorganisms.

Impact of blood collection devices on clinical chemistry assays.

Blood collection devices interact with blood to alter blood composition, serum, or plasma fractions and in some cases adversely affect laboratory tests. Vascular access devices may release coating substances and exert shear forces that lyse cells. Blood-dissolving tube additives can affect blood constituent stability and analytical systems. Blood tube stoppers, stopper lubricants, tube walls, surfactants, clot activators, and separator gels may add materials, adsorb blood components, or interact with protein and cellular components. Thus, collection devices can be a major source of preanalytical error in laboratory testing. Device manufacturers, laboratory test vendors, and clinical laboratory personnel must understand these interactions as potential sources of error during preanalytical laboratory testing. Although the effects of endogenous blood substances have received attention, the effects of exogenous substances on assay results have not been well described. This review will identify sources of exogenous substances in blood specimens and propose methods to minimize their impact on clinical chemistry assays.

Improved detection of intact tyrosine sulfate-containing peptides by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry in linear negative ion mode.

Sulfation of tyrosine residues is a common post-translational modification, but detecting and quantitating this modification poses challenges due to lability of the sulfate group. The goal of our studies was to determine how best to detect and to assess the stoichiometry of this modification using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOF MS). Sulfated and nonsulfated forms of peptides-hirudin(55-65), caerulein, and cholecystokinin octapeptide and phosphorylated and nonphosphorylated pp60-c-src (521-533)-were analyzed using several matrices: sinapinic acid (SA), 2,5-dihydroxybenzoic acid (DBA), and cyano-4-hydroxycinnamic acid (CHCA). Intact sulfated peptides were difficult to detect using positive ion mode; peptides were observed as desulfated ions. Phosphorylated peptide was stable and was detected in positive and negative ion modes. Detection of sulfated peptides improved with: (1) Analysis in negative ion mode, (2) Decreased laser power, (3) Matrix selection: DBA>/=SA>CHCA. In negative ion mode, desorption/ionization of sulfated peptide was equivalent or more efficient than nonsulfated peptide, depending on conditions of analysis. Examination of a tryptic digest of alpha(2)-antiplasmin detected the single site of sulfation in negative ion mode but not in positive ion mode. We conclude that improved detection of sulfated peptides can be achieved in negative ion mode. Dual analysis in positive and negative ion modes serves as a potential means of identifying peptides with labile modifications such as sulfation and distinguishing them from phosphorylation.

The dynamic range problem in the analysis of the plasma proteome.

One of the greatest challenges in analyzing the plasma proteome is the wide range of concentration of different proteins. The current study examines the range of protein concentration for 18 proteins measured over a year in a clinical laboratory to provide data on pathological extremes in protein concentrations. The complete measured range, from upper limits for albumin to lowest values for thyroid-stimulating hormone (TSH), represented more than 10 logs of molar abundance. A number of plasma proteins measured in the clinical laboratory varied over a concentration range spanning more than 4 logs, and limits of detection of clinical assays were inadequate to assess full concentration ranges of several proteins. Considering reported values from studies using higher sensitivity assays suggest that plasma concentrations of prostate-specific antigen (PSA), human chorionic gonadotropin (hCG), and cardiac troponin I vary by more than 7 logs. All of the plasma proteins measured in the present study represent secretory proteins or highly expressed components of specific tissues. Thus, the dynamic range for these components is likely to greatly underestimate the total range of protein concentration in the plasma proteome.

Comparison of urinary albumin quantification by immunoturbidimetry, competitive immunoassay, and protein-cleavage liquid chromatography-tandem mass spectrometry.

BACKGROUND: Increased urinary albumin excretion is a well-documented diagnostic and prognostic biomarker for renal disease. Urinary albumin is typically measured in clinical settings by immunoassay methods. However, neither a reference method nor a urine albumin calibration reference material is currently available. METHODS: We quantified urinary albumin in patient samples by using 3 commercially available reagent systems: DiaSorin SPQ and Beckman Coulter LX 20 (immunoturbidimetric), and Siemens Immulite (competitive immunoassay). Results were compared to values obtained by protein-cleavage liquid chromatography-tandem mass spectrometry (LC-MS/MS). RESULTS: In general, results from the 3 immunoassays agreed with results from LC-MS/MS. However, the SPQ results showed a negative bias across all ranges of albuminuria [(0-200 mg/L, y = 0.91x - 3.74 (CI 0.86-0.96); > 200 mg/L, y = 0.88x - 40.30 (CI 0.76-1.00)], whereas the LX 20 showed minimal bias in the 0-200 mg/L range [y = 0.97x - 88 (CI 0.92-1.02)] and the Immulite assay showed positive bias in the 0-200 mg/L range [y = 1.15x - 4.38 (CI 1.09-1.20)]. CONCLUSIONS: These results showed a reasonable quantification of urinary albumin by representative polyclonal and monoclonal immunoassays compared to an LC-MS/MS assay. In addition, the results do not suggest the presence of nonimmunoreactive albumin in urine. However, differences in analytic performance between assays support the need for a reference calibration material and reference method to standardize clinical laboratory measurements of urinary albumin.

Proteinuria without albuminuria: urinary protein excretion by a subset of patients with burn injuries.

BACKGROUND: There is disagreement regarding the utility of urinary albumin excretion as a marker for capillary injury in patients with severe burn injuries. We examined protein components in urine specimens from patients with burn injury. METHODS: Detailed analysis was performed for a set of 5 urine specimens selected based on a high ratio of albumin-sized molecules by size-exclusion HPLC (Accumin) versus albumin by immunoassay methods. Specimens were analyzed for total protein, alpha(1)-microglobulin, alpha(1)-acid glycoprotein, cystatin C, and retinol-binding protein. Urine components were analyzed by chromatographic and electrophoretic methods. Major components were identified by mass spectrometry of tryptic peptides. RESULTS: A subset of urine specimens had increased total protein with slight increases in albumin by immunoassay or by polyacrylamide gel electrophoresis. Albumin values by size-exclusion HPLC were more than 10-fold higher. Immunoassays for alpha(1)-microglobulin and alpha(1)-acid glycoprotein yielded concentrations 5-10 fold higher than for albumin. Other major components identified included zinc-alpha(2)-glycoprotein and leucine-rich-alpha(2)-glycoprotein. CONCLUSIONS: A subset of patients with burn injury had increased total urinary protein resulting primarily from increased excretion of proteins such as alpha(1)-microglobulin and alpha(1)-acid glycoprotein with little increase in albumin excretion. The unusual composition of urinary proteins in these patients may relate to decreased filtered load of albumin and increased filtered load of acute phase reactants or to alterations in renal tubular protein processing. Thus, measurement of urinary albumin may have decreased sensitivity for detecting kidney injury in burn patients.

Urine albumin measurement: effects of urine matrix constituents.

BACKGROUND: There is increasing clinical interest in accurate measurement of urine albumin as an early indicator of kidney disease. However, urine is a highly variable sample matrix that may exert matrix effects on assays for urine albumin. METHODS: Variation in urine composition was assessed for components routinely measured in the clinical laboratory over 1 year. Solutions of representing different concentrations of urine components were prepared and spiked with a constant amount of albumin. Fourteen urine specimens of variable composition were ultrafiltered and spiked with a constant amount of albumin. Concentrations of albumin in solutions were determined with an immunoturbidimetric assay on the Beckman LX20 analyzer and with a competitive immunoassay on the Siemens Immulite analyzer. RESULTS: Variation in concentrations of most constituents altered measured urine by <10%. High NaCl concentration increased assay values by 10% or more. Addition of 0.5% Triton X-100 increased values in the LX20 assay by about 20%. Most ultrafiltered urine specimens had <10% effect on measured albumin at a concentration near 20 mg/l, but one specimen was noted to decrease measured albumin by approximately 20% in the LX20 assay. CONCLUSIONS: Urine matrix components exert small but potentially significant effects on assays for urine albumin. Urine matrix effects should be considered in the design and evaluation of assays for albumin.

Current issues in measurement and reporting of urinary albumin excretion.

BACKGROUND: Urinary excretion of albumin indicates kidney damage and is recognized as a risk factor for progression of kidney disease and cardiovascular disease. The role of urinary albumin measurements has focused attention on the clinical need for accurate and clearly reported results. The National Kidney Disease Education Program and the IFCC convened a conference to assess the current state of preanalytical, analytical, and postanalytical issues affecting urine albumin measurements and to identify areas needing improvement. CONTENT: The chemistry of albumin in urine is incompletely understood. Current guidelines recommend the use of the albumin/creatinine ratio (ACR) as a surrogate for the error-prone collection of timed urine samples. Although ACR results are affected by patient preparation and time of day of sample collection, neither is standardized. Considerable intermethod differences have been reported for both albumin and creatinine measurement, but trueness is unknown because there are no reference measurement procedures for albumin and no reference materials for either analyte in urine. The recommended reference intervals for the ACR do not take into account the large intergroup differences in creatinine excretion (e.g., related to differences in age, sex, and ethnicity) nor the continuous increase in risk related to albumin excretion. DISCUSSION: Clinical needs have been identified for standardization of (a) urine collection methods, (b) urine albumin and creatinine measurements based on a complete reference system, (c) reporting of test results, and (d) reference intervals for the ACR.

High-abundance polypeptides of the human plasma proteome comprising the top 4 logs of polypeptide abundance.

BACKGROUND: Plasma contains thousands of proteins, but a small number of these proteins comprise the majority of protein molecules and mass. CONTENT: We surveyed proteomic studies to identify candidates for high-abundance polypeptide chains. We searched the literature for information on the plasma concentrations of the most abundant components in healthy adults and for the molecular mass of the mature polypeptide chains in plasma. Because proteomic studies usually dissociate proteins into polypeptide chains or detect short peptide segments of proteins, we summarized data on individual peptide chains for proteins containing multiple subunits or polypeptides. We collected data on about 150 of the most abundant polypeptides in plasma. The abundant polypeptides span approximately the top 4 logs of concentration in plasma, from 650 to 0.06 micromol/L on a molar basis or from about 50,000 to 1 mg/L mass abundance. CONCLUSIONS: Data on the concentrations of the high-abundance peptide chains in plasma assist in understanding the composition of plasma and potential approaches for clinical laboratory or proteomic analysis of plasma proteins. Development of more extensive databases regarding the plasma concentrations of proteins in health and diseases would promote diagnostic and proteomic advances.

Analysis of molecular forms of albumin in urine.

Albumin is not only one of the most abundant urinary protein components and one of the most widely used clinical markers for kidney disease, it also is a significant source of molecular complexity of the urinary proteome and peptidome. Urine contains multiple fragments and modified forms of albumin. Analysis of molecular forms of albumin in urine is of fundamental importance for understanding clinical assays for albumin quantification, specimen stability, and proteomic and peptidomic analysis of urine.

Pseudohyperphosphatemia associated with high-dose liposomal amphotericin B therapy.

BACKGROUND: Acute increases in serum inorganic phosphorus (Pi) up to 4.75 mmol/l in the absence of hypocalcemia and tissue deposition of calcium phosphate were noted in 3 patients receiving liposomal amphotericin B (L-AMB). We investigated L-AMB as a possible cause of pseudohyperphosphatemia. METHODS: Serum samples from the index patient were analyzed for Pi content by our laboratory's primary analyzer (Synchron LX20) and by an alternate analyzer (Vitros). Clear and lipemic serum pools, and normal saline, were spiked with L-AMB and analyzed by the LX20 Pi method. Ultrafiltration studies were performed on patient and spiked sera. RESULTS: Increased Pi values were obtained only from the LX20 analyzer. There was a direct linear relationship between the concentration of L-AMB in the spiked samples and the LX20 Pi results, indicating a 0.9 mmol/l Pi increase for every 100 mg/l increase in L-AMB. Ultrafiltration normalized the Pi results. CONCLUSION: Serum Pi results may be falsely increased in patients receiving L-AMB when measured by the LX20 analyzer. This novel cause of pseudohyperphosphatemia is due to interference of L-AMB with the method and is corrected by ultrafiltration of the specimen. Since the LX20 analyzer is widely used by the clinical laboratories clinicians and laboratory personnel should recognize this interference in order to avoid unnecessary diagnostic procedures and interventions.