Analysis of low-angle x-ray scattering peaks from lyophilized biological samples

Low-angle x-ray scattering (LAXS) from lyophilized blood and its constituents is characterized by the presence of two peaks in the forward direction of scattering. These peaks are found to be sensitive to the variations in the molecular structure of a given sample. The present work aims to explore the nature of LAXS from a variety of lyophilized biological samples. It also aims to investigate the possibility that a certain biological macromolecule is responsible of the production of LAXS peaks. This is carried out through measurements of LAXS from complex biological samples and their basic constituents. Among the measured samples are haemoglobin (Hb), globin, haem, packed red blood cells, bovine albumin, egg albumin, milk, casein, glutamine, alanine, fat, muscle and DNA. A table containing some characteristic parameters of the LAXS profiles of these samples is also presented. Analysis of measured profiles shows that all lyophilized samples produce at least one relatively broad peak at a scattering angle around 10.35 degrees. The full width at half maximum (FWHM) of this peak varies considerably among the measured samples. Except for milk and casein. one additional peak at a scattering angle around 4.65 degrees is observed only in the LAXS profiles of proteins or protein-rich samples. This fact strongly suggests protein to be the biological macromolecule from which this characteristic peak originates. The same idea is further strengthened through discussion of some previous observations.     

The diagnostic capability of x-ray scattering parameters for the characterization of breast cancer

PURPOSE: The evaluation of the diagnostic capability of easy to measure x-ray scattering profile characterization parameters for the detection of breast cancer in excised samples. The selected parameters are the full width at half maximum (FWHM) and area under the x-ray scattering profile of breast tissue in addition to the ratio of scattering intensities (I2/I1%) at 1.6 nm(-1) to that at 1.1 nm(-1) (corresponding to scattering from soft and adipose tissues, respectively). Methods: A histopathologist is asked to classify 36 excised breast tissue samples into healthy or malignant. A conventional x-ray diffractometer is used to acquire the scattering profiles of the investigated samples. The values of three profile characterization parameters are calculated and the diagnostic capability of each is evaluated by determining the optimal cutoffs of scatter diagrams, calculating the diagnostic indices, and plotting the receiver operating characteristic (ROC) curves. Results: At the calculated optimal cutoff for each of the examined parameters, the sensitivity ranged from 78% (for area under curve) up to 94% (for FWHM), the specificity ranged from 94% [for I2/I1% and area under curve] up to 100% (for FWHM), and the diagnostic accuracy ranged from 86% (for area under curve) up to 97% (for FWHM). The area under the ROC curves is greater than 0.95 for all of the investigated parameters, reflecting a highly accurate diagnostic performance. Conclusions: The discussed tests offered a means to quantitatively evaluate the performance of the suggested breast tissue x-ray scattering characterization parameters. The performance results are promising, indicating that the evaluated parameters would be considered a tool for fast, on spot probing of breast cancer in excised tissue samples.     

Characterization of cirrhosis and hepatocellular carcinoma using low-angle x-ray scattering signatures of serum

The diagnosis of hepatocellular carcinoma (HCC) usually occurs at late stages in the disease when there are few effective treatment options. The measurement of the concentration of tumour markers in the serum of patients is a complementary tool frequently used for the interpretation of diagnostic imaging results. It is also used as a prognostic tool for the detection of cancer. Unfortunately, the sensitivity of tumour markers is still low and many times it yields normal results for cirrhotic and HCC patients. In the current work, the detection possibility of the structural changes in serum proteins accompanying cirrhosis and HCC is investigated using a low-angle x-ray scattering (LAXS) technique. The results show that there are significant differences in the LAXS profiles of cirrhosis and HCC lyophilized serum samples compared to normal. The changes in shape, total counts and position of the first scattering peak at 4.8 degrees, which was previously reported to be sensitive to the structural changes in protein, showed the most characteristic deviations from normal serum. The present results are promising and would offer a potentially helpful complementary tool for monitoring cirrhosis and HCC.     

Characteristics of low-angle x-ray scattering from some biological samples

Design of medical imaging devices based on the detection of low-angle coherent scattering is a subject of increasing interest. The technique is based on the differences in the distribution of photons coherently scattered from different body tissues. Coherent scattering is also useful in monitoring changes that may occur in a healthy tissue (e.g. carcinoma). In this work, low angle scattering properties of some tissues and tissue-equivalent materials are studied. Special care is given to the possibility of distinguishing between tissues of similar water content (e.g. muscle and blood). For this purpose, a Monte Carlo simulation is updated, introducing molecular form factor data, which include molecular interference effects. This program is used to simulate the angular distribution of scattered photons from two tissue-equivalent materials (lucite and water) and three biological samples (muscle, fat and blood). Simulation results agree well with previously measured angular distributions of scattered photons at 59.54 keV. Scattering from water and lucite is also measured at 8.047 keV. The effects of scattering geometry, sample thickness, incident photon energy and tissue type on the angular distribution of scattered photons are investigated. Results reveal the potential of measuring the full width at half maximum (FWHM) of the scattered photon distribution for tissue characterization. Energies up to 13 keV and sample thickness of 0.3 cm reported maximum differences between investigated samples. These conditions are expected to maximize the potential of using coherent scattering set-ups to monitor changes in biological samples even if their water contents are similar. Present results may act as a guide for the optimization of coherent scattering imaging systems.     

650nm低能量激光照射离体高血脂全血和红细胞对其吸收光谱的影响

目的 研究低能量激光照射离体高血脂全血和红细胞对其吸收光谱的影响.方法 选取30例高血脂患者血样为高血脂组,30例正常血样为正常组,用多功能酶标仪分别对血样的全血和红细胞进行扫描,比较两个样本吸收光谱的异同;选用低能量650 nm激光照射待测血样,用多功能酶标仪对全血和红细胞进行扫描并比较照射前后吸收光谱的变化.结果 高血脂血样的全血和红细胞在416、544、578 nm处的吸收峰值均比正常血样高,因此,可通过光谱吸收峰初步判定血液中血脂异常的情况.650 nm激光照射30例高血脂血样后,红细胞在416、544、578 nm处峰值分别降低,照射前后差异具有统计学意义（P＜0.05）;全血照射后其吸收光谱在416、544、578 nm升高,差异具有统计学意义（P＜0.05）.结论 650 nm低能量激光照射可降低高血脂红细胞对光谱的吸收峰,且有助于改善红细胞膜-浆脂质的平衡,从光谱学角度证明了激光治疗高血脂疾病的可行性.     

Interference of hemoglobin Hope on beta-thalassemia diagnosis by the capillary electrophoresis Method

The β-chain hemoglobin (Hb) variants interfere with the diagnosis of β-thalassemia trait using high-performance liquid chromatography (HPLC) and capillary electrophoresis (CE). We analyzed the effect of Hb Hope, a β-chain Hb variant frequently found in the Thai population, on β-thalassemia trait diagnosis. HPLC and CE were used to quantify the level of HbA(2) in 11 whole blood samples containing Hb Hope. The levels of Hb Hope detected by both methods were similar. An elevated HbA(2) level was found in all samples analyzed by the CE method, while 1 was increased when analyzed by HPLC, which was a compound heterozygous of Hb Hope and α-thalassemia-1 SEA-type deletion. Of 11 samples, 6 had mean corpuscular volumes within the reference range. All samples showed negative results for molecular analysis of β(0)-thalassemia codon 17, 41/42, and 71/72 mutations and β-thalassemia 3.5-kb deletion. Therefore, Hb Hope interfered with the diagnosis of β-thalassemia trait analyzed by CE but not by HPLC.     

Changes in serum lipid concentrations during iron depletion and after iron supplementation

To investigate the effects of body iron depletion and iron supplementation on serum lipid concentrations, hematologic indices, iron markers, and serum lipid profiles were measured in 427 girls, age 14-19 yr. There were no significant differences in serum lipid concentrations between subjects with moderate iron deficiency anemia (blood Hb < 12.0 g/dL) and healthy controls. However, serum total cholesterol concentration (mean +/- SD, 148 +/- 16 mg/dL) in severely anemic subjects with blood Hb < 8.0 g/dL was significantly lower than in subjects with blood Hb > or = 14.0 g/dL (170 +/- 17 mg/dL) (p < 0.01). Moreover, serum triglyceride concentration in subjects with blood Hb > 14.0 g/dL was 2-fold higher than in the severely anemic subjects. Mean values of serum total cholesterol and triglyceride (149 +/- 17 mg/dL and 58 +/- 22 mg/dL) in girls with severe anemia were significantly elevated after iron supplementation (164 +/- 17 mg/dL and 98 +/- 26 mg/dL) (p < 0.01, respectively). In the severely anemic subjects, blood Hb concentration was correlated with serum total cholesterol (r = 0.49, p < 0.01) and triglyceride concentrations (r = 0.51, p < 0.01). These findings indicate that severe iron deficiency anemia in girls is attended by decreased concentrations of serum total cholesterol and triglyceride, and that these reduced serum lipid levels return to normal following iron supplementation.     

X-ray scattering for classifying tissue types associated with breast disease

Collagen types I and III can be characterized at the molecular level (at the tens to hundreds of nanometers scale) using small angle x-ray scattering (SAXS). Although collagen fibril structural parameters at this length scale have shown differences between diseased and nondiseased breast tissues, a comprehensive analysis involving a multitude of features with a large (>50) patient cohort has not previously been investigated. Breast tissue samples were excised from 80 patients presenting with either a breast lump or reduction mammoplasty. From these, invasive carcinoma, benign tissue, and normal parenchyma were analyzed. Parameters related to collagen structure, including longitudinal (axial) and lateral (equatorial) features, polar angle features, total scattering intensity, and tissue heterogeneity effects, were extracted from the SAXS patterns and examined. The amplitude of the third-order axial peak and the total scattering intensity (amorphous scatter) showed the most separation between tissue groups and a classification model using these two parameters demonstrated an accuracy of over 95% between invasive carcinoma and mammoplasty patients. Normal tissue taken from disease-free patients (mammoplasty) and normal tissue taken from patients with presence of disease showed significant differences, suggesting that SAXS may provide different diagnostic information from that of conventional histopathology.     

Postmortem blood ferritin concentrations in sudden infant death syndrome.

AIMS--To confirm the observation of extremely high concentrations of ferritin in postmortem serum samples in sudden infant death syndrome (SIDS); to examine the factors influencing blood ferritin concentrations postmortem; to determine whether or not these high blood ferritin concentrations are characteristic of SIDS. METHODS--Postmortem samples of cardiac blood were obtained from 58 full term infants who died of SIDS and 14 full-term infants who died of a variety of other causes. Whole blood and serum ferritin concentrations were determined and compared with age at death, liver iron concentration, serum iron concentration, and serum lactate dehydrogenase activity. RESULTS--The median postmortem blood ferritin concentration for all infants was 18,600 micrograms/l, which is about 200 times the concentration found in the serum of normal, live infants. Serum iron concentrations were high and there was a highly significant correlation between serum ferritin and iron concentrations suggesting that much of the serum iron was contributed by ferritin. There was no significant difference between serum and whole blood ferritin concentrations. H to L type ferritin ratios were higher in blood from the left than the right ventricle of the heart but the ferritin was always predominantly L type. Blood ferritin concentrations rose rapidly after death but in samples collected at postmortem examination there was a significant correlation with liver iron concentration and an inverse correlation with age. Median values for blood ferritin were higher in SIDS (22,500; n = 58) than in control cases (6900; n = 7) dying under one year of age; however, in both groups ferritin concentrations decreased with age. CONCLUSIONS--Release of ferritin into the blood postmortem seems to be characteristic of infants dying before the age of one year rather than characteristic of SIDS. Two factors may cause such ferritin release postmortem: tissue breakdown and the high level of storage iron in cells of the reticuloendothelial system (including endothelial cells lining vessel walls). SIDS occurs when tissue iron concentrations are higher than at any other time of life. It is possible that the ready availability of iron enhances free radical damage which might be implicated in SIDS.     

Conjugate of bovine hemoglobin and human serum albumin as a candidate for blood substitute: characteristics and effects on rats

Conjugate of bovine hemoglobin (bHb) and human serum albumin (HSA) was prepared. The product was simply composed of 89.7% one-to-one Hb-HSA conjugate, 6.0% oligomer of Hb and HSA, 3.5% unconjugated HSA and 0.8% unconjugated Hb, with an average molecular weight of 157 kD. The physicochemical characteristics were determined. Effects of single replacement on blood pressure and long-term survival of rats with 30% and 60% acute blood loss were studied, in comparison with Ringer-lactate solution, stroma-free hemoglobin (SFHb), 5% HSA in Ringer-lactate, whole blood and no resuscitation fluid. Results showed that Hb-HSA conjugate maintained the mean arterial pressure of rats to initial level with no pressor effect. Long-term effects of the replacement fluids on 30% bleeding rats showed that, for the group infused with Hb-HSA conjugate, histology of five major organs, heart, kidney, liver, spleen and lung, were essentially normal, similar to that of whole blood, while obviously renal side-effects appeared in other groups. The efficacy of the conjugate was further demonstrated by the resuscitation of lethal hemorrhagic shock rats (60% acute blood loss) with 100% survival rate (followed for 14 days), the same result as whole blood. The Hb-HSA conjugate can thus be another candidate for blood substitute in emergency.     

Study of bovine hemoglobin dissociation by multiangle laser light-scattering method

Hemoglobin (Hb) is an oligomeric protein, composed of four monomeric subunits. Hb molecule may undergo dissociation from a single native tetramer to two dimmers, which is called hemoglobin dissociation. In this article the dissociation of bovine Hb is studied by measurment of the average MW of the samples using the multiangle laser light-scattering method. Advanced multiangle laser light-scattering technique is a powerful method to determine the absolute molecular weights of the protein in solution. Two different methods, microbatch multiangle light-scattering (MALS) and on-line size-exclusion high-performance liquid chromatography light scattering with refractive index detector, are used to measure the average molecular weight of bovine Hb in different concentration respectively. The results of the two methods are agreed well. From the results, it can be concluded that the average molecular weigh of bovine Hb will be about 54 kDa when the bovine Hb concentration is more than 1.5 mg/mL, and will be about 36 kDa when the concentration is less than 0.03 mg/mL. The other conclusion, which can be derived from these results, is that the dissociation of bovine Hb is related with the pH and the tetramer appears to be more stable in the pH range of 6-9.     

Evaluation of 24-second cyanide-containing and cyanide-free methods for whole blood hemoglobin on the Technicon H*1TM analyzer with normal and abnormal blood samples

Automated methods, with and without cyanide (+CN and -CN), for whole blood hemoglobin (Hb) determination were evaluated on the Technicon H*1TM System. Both automated Hb methods were linear over the range 0-250 g/L (0-25 g/dL) and correlated well with the International Committee for Standardization of Hematology (ICSH) reference method and with the Coulter S+II. Both methods quantitatively converted whole blood containing up to 100% carboxyhemoglobin in less than 24 seconds to their respective end products. With respect to abnormal samples (sickle cell anemia, multiple myeloma, and hyperlipemia), both H*1 methods gave Hb results that were equivalent to the (postfiltration) ICSH method. For samples with white blood cell (WBC) counts less than 36 X 10(9)/L, the +CN method was equivalent to the (postfiltration) ICSH method, whereas for WBC counts greater than 20 X 10(9)/L, the -CN method showed acceptable recovery of the mean but unacceptable imprecision. For WBC counts of 36-164 X 10(9)/L, the +CN method yielded acceptable Hb recovery with unacceptable imprecision. Hyperlipemia, resulting from addition of Intralipid directly to the blood samples, caused large errors in both H*1 methods.     

Trace element analysis in biological samples by using XRF spectrometry with secondary radiation

An x-ray fluorescence method for in vitro analysis of trace elements in biological samples is presented. The method is characterised by the use of an x-ray tube with secondary targets as a monoenergetic radiation source, and by 'infinitely thin' specimens. In the experimental work, different aspects have been examined in order to optimise the sensitivity of the method. It is shown that it is extremely important to use collimators of high purity materials and very pure and thin sample supports. Regarding the geometry, it is pointed out that a collimator between specimen and detector reduces the counting rate caused by scattering in air and other materials. Scattering in the biological matrix is reduced by preconcentration of biological liquids or tissues. The method has been applied to the analysis of blood serum samples. Typical sensitivities for Fe and Rb are 1.6 ng cm-2 and 1.5 ng cm-2 respectively, in a counting time of 10(3) s.     

Serum erythropoietin and interleukin-6 levels in hemodialysis patients with hepatitis virus infection.

The influence of hepatitis B (HBV) and hepatitis C virus (HCV) infection on blood hemoglobin (Hb) and serum erythropoietin (Epo) and interleukin-6 (IL-6) concentrations was studied in 48 anemic patients on regular hemodialysis. They were grouped as follows: (I) 19 patients whose Hb values improved after infection (Hb > 85 g/L), (II) 10 patients with persisting anemia after infection (Hb < 75 g/L), and, without hepatitis virus markers (III) 8 patients with Hb > 85 g/L and (IV) 11 patients with Hb < 75 g/L. Serum immunoreactive Epo levels were significantly higher in group I (34.4+/-47.1 U/L) than in the other groups (II, 10.8+/-6.0; III, 7.9+/-3.2; IV, 8.4+/-4.3). Serum IL-6 was higher in group I than group III (7.7+/-7.8 pg/ml vs. 3.6+/-2.4; p = 0.05) but similar to the other groups. Hb levels in group I were maximal at the time of serum alanine aminotransferase normalization. Red cell production increases as a result of elevated circulating Epo during hepatic regeneration after HBV or HCV infection.     

Spectroscopic quantitative analysis of blood glucose by Fourier transform infrared spectroscopy with an attenuated total reflection prism.

As an alternative way of long-term glycemic monitoring, the glucose measurement by analyzing Fourier transform infrared absorbance spectra with an attenuated total reflection prism has been developed. In glucose aqueous solution, glucose has characteristic absorptions at the wave numbers of 1033 and 1080 cm-1 and the absorption intensities are proportional to glucose concentrations. In serum and whole blood samples, however, red blood cell corpuscles, serum albumin and serum gamma-globulin interfere with the absorbance spectra of glucose and shift the base line upward significantly. Therefore, to eliminate these interferences in serum and whole blood samples, the feasibility of the calibration curves obtained by using difference absorbance spectra with those of fasting samples was studied. As a result, highly significant correlations between glucose concentrations estimated by Fourier transform infrared spectroscopic method and those measured by glucose oxidase method were obtained (r = 0.981 and 0.989 for serum and whole blood samples, respectively). From these experiments, it was concluded that by infrared spectroscopy glucose concentrations in the serum and whole blood samples could be measured quantitatively or monitored if the base line drifts and interferences were subtracted.     

The small-animal radiation research platform (SARRP): dosimetry of a focused lens system

A small animal radiation platform equipped with on-board cone-beam CT and conformal irradiation capabilities is being constructed for translational research. To achieve highly localized dose delivery, an x-ray lens is used to focus the broad beam from a 225 kVp x-ray tube down to a beam with a full width half maximum (FWHM) of approximately 1.5 mm in the energy range 40-80 keV. Here, we report on the dosimetric characteristics of the focused beam from the x-ray lens subsystem for high-resolution dose delivery. Using the metric of the average dose within a 1.5 mm diameter area, the dose rates at a source-to-surface distance (SSD) of 34 cm are 259 and 172 cGy min(-1) at 6 mm and 2 cm depths, respectively, with an estimated uncertainty of +/-5%. The per cent depth dose is approximately 56% at 2 cm depth for a beam at 34 cm SSD.     

血液透析联合血液灌流治疗尿毒症皮肤瘙痒症的疗效

目的 观察尿毒症皮肤瘙痒症（UP）患者在血液透析（HD）联合血液灌流（HP）治疗的疗果。方法 选取2016年6月~2017年6月雅安市第二人民医院血液透析中心诊断为UP患者40例,行HD联合HP治疗,共6个月。比较患者治疗前后四项目瘙痒（Four-item Itch Questionnaire,FIIQ）量表评分、血白细胞（WBC）、全血红细胞（Hb）、血清肌酐（Cr）、血清尿素氮（Bun）、胱抑素C（Cys-C）、血清磷（P）、β2-微球蛋白（β2-MG）、甲状旁腺素（PTH）、C反应蛋白（CRP）、白细胞介素6（IL-6）、肿瘤坏死因子α（TNF-α）水平。结果 HD联合HP治疗6个月后,36例（4例排除）患者WBC、Hb水平与治疗前比较,差异无统计学意义（P＞0.05）;治疗后FIIQ量表评分、Cr、BUN、Cys-C、P、β2-MG、PTH、CRP、IL-6、TNF-α较治疗前降低,差异有统计学意义（P＜0.05）。结论 HD联合HP有助于更好的清除UP患者体内的中小分子物质、调节水、电解质平衡,缓解UP患者的症状,值得借鉴。     

Measurement of CT radiation profile width using CR imaging plates

This paper describes the procedure for using a Fuji computed radiography (CR) imaging plate (IP) for the measurement of computed tomography (CT) radiation profiles. Two sources of saturation in the data from the IP, signal and quantization, were characterized to establish appropriate exposure and processing conditions for accurate measurements. The IP generated similar profiles compared to those obtained from digitized ready-pack films, except at the profile edges, where the exposure level is low. However, when IP pixel values are converted to exposure, CR and digitized film profiles are in agreement. The full width at half maximum (FWHM) of the CT radiation profile was determined from the relationship between pixel value and exposure and compared to FWHM of the digitized optical density profile from film. To estimate the effect of scattering by the cassette material, radiation profiles were acquired from IPs enclosed in a cassette or in a paper envelope. The presence of the cassette made no difference in the value determined for FWHM. With proper exposure and processing conditions, the FWHM of 5, 10, and 15 mm collimated beams were measured using IPs to be 7.1, 11.9, and 17.0 mm and using film to be 7.2, 12.2, and 16.8 mm, respectively. Our results suggest that, under appropriate conditions, the estimation of the width of the CT radiation profile using Fuji CR is comparable to the measurement from film density described in American Association of Physicists in Medicine (AAPM) Report No. 39. Although our experiment was conducted using Fuji CR, we anticipate that CR plates from other vendors could be successfully used to measure CT beam profiles because of similar empirical relationships between pixel value and exposure.     

Use of a blood glucose meter for radiochromic film analysis in blood irradiation

The use of a diabetic blood glucose meter for radiochromic film dosimetry in blood irradiation using x-ray beams on a medical linear accelerator has been investigated. The glucose meter provides optical density analysis in the visible and infrared region using a reflectance measurement technique. By comparing the 'blood sugar' level output with standard calibration gafchromic films a calibration curve is produced for quantitative analysis. Results show that a reproducible dose to meter output curve can be fitted using a second order polynomial function and that blood irradiation doses in vitro were measured to within 7.9% mean error (as compared to ionization chamber results) using the blood glucose meter. This level of accuracy falls below that measured with a standard densitometer (4.3%); however, results show that the blood glucose meter, which would be available in any haematology department, produces an adequate measure of gafchromic film optical density for blood irradiation dosimetry.     

"M" alpha-1-antitrypsin at term of pregnancy in an African population: concentration and microheterogeneity

For 19 multipars in Ivory Coast alpha-1-antitrypsin (A1AT) has been determined in mother's serum as well as in the cord serum and the amniotic fluid. In all cases the type of A1AT was MM. In 15 cases the pregnancies were normal and full term while four were premature (28th week). The A1AT levels obtained were found to be similar to the literature values and thereby do not seem to be influenced by the ethnic origin. Crossed electroimmunodiffusion analyses demonstrate a distribution of the different fractions of A1AT which is characteristic for each of the above groups of samples analysed. The similarity of the fractions in mother's serum and amniotic fluid indicate a maternal origin for A1AT of the latter. The cord blood shows a very characteristic increase in the peak M6. In the author's experience, they have only noticed such a peak in the blood of patients with primary liver cancer of the phenotype MM.