Delta Bilirubin Research Articles

Updating Pediatric Reference Ranges at Seattle Children’s Hospital Using an Indirect Approach

Abstract Background The aim of this project was to evaluate the accuracy of historical reference ranges at our institution. Our population of interest is children (<18 years old) within a large city. Ideally, reference intervals are determined using the direct method by performing studies on healthy volunteers. However, this approach has many challenges including being resource intensive, and, particularly in the pediatric population, difficulty in obtaining the required number of healthy volunteers for every age range. Given these challenges, a second approach, the indirect approach, has also been utilized to update reference ranges. This approach uses retrospective data analysis on data that was not specifically collected for evaluation of a reference range. Abnormal values are filtered out using a variety of methods. Methods For our project we pulled all outpatient data between 10/3/2020 and 06/30/2023 for the following analytes: albumin, ALP, AST, ALT, BUN, calcium, chloride, CO2, conjugated bilirubin, delta bilirubin, potassium, sodium, total bilirubin, total protein, and unconjugated bilirubin. We limited each patient to their first result for each analyte on the basis that analytes being measured more than once are more likely to be abnormal, and to limit bias. We also pulled the age (in days) and legal sex for each patient. The Tukey method was used to exclude outliers on the assumption that most data points would be normal, so any outliers would represent abnormal values from potentially not healthy patients. For each analyte and every age range the frequency of each value was plotted and the percentage of values within the current reference range, 2.5th percentile, and 97.5th percentile were calculated. A reference range was considered accurately representative of the normal population at our institution if the percentage of values within the reference range was greater than or equal to 80%. Results The “in range” percentage for albumin, ALT, BUN, calcium, chloride, conjugated bilirubin, delta bilirubin, creatinine, and total bilirubin was greater than 80% for all age ranges tested. Potassium, sodium, and unconjugated bilirubin each had one age range where the percent “in range” was less than 80% (2 days – 3 months, < 1 week, and < 1 month respectively), however the sample number for these age ranges was considered too low to make any significant conclusions (n=273, n=11, and n=21 respectively). ALP, AST, CO2, and total protein each had at least one age range where the percent “in range” was less than 80% and a substantial number of data points was available. In these cases, we proposed changing the reference interval to the calculated 2.5th and 97.5th percentiles based on our retrospective data analysis or verifying an externally validated pediatric reference interval from the CALIPER study.

Development of an equation to predict delta bilirubin levels using machine learning

ObjectiveDelta bilirubin (albumin-covalently bound bilirubin) may provide important clinical utility in identifying impaired hepatic excretion of conjugated bilirubin, but it cannot be measured in real-time for diagnostic purposes in clinical laboratories. MethodsA total of 210 samples were collected, and their delta bilirubin levels were measured four times using high-performance liquid chromatography. Data collected included age, sex, diagnosis code, delta bilirubin, total bilirubin, direct bilirubin, total protein, albumin, globulin, aspartate aminotransferase, alanine transaminase, alkaline phosphatase, gamma-glutamyl transferase, lactate dehydrogenase, hemoglobin, serum hemolysis value, hemolysis index, icterus value (Iv), icterus index (Ii), lipemia value (Lv), and lipemia index. To conduct feature selection and identify the optimal combination of variables, linear regression machine learning was performed 1,000 times. ResultsThe selected variables were total bilirubin, direct bilirubin, total protein, albumin, hemoglobin, Iv, Ii, and Lv. The best predictive performance for high delta bilirubin concentrations was achieved with the combination of albumin-direct bilirubin-hemoglobin-Iv-Lv. The final equation composed of these variables was as follows: delta bilirubin = 0.35 × Iv + 0.05 × Lv − 0.23 × direct bilirubin − 0.05 × hemoglobin − 0.04 × albumin + 0.10. ConclusionThe equation established in this study is practical and can be easily applied in real-time in clinical laboratories.

Measurement and Clinical Usefulness of Delta Bilirubin in Liver Disease

Delta bilirubin is albumin bound conjugated bilirubin that can be calculated by formula [Total bilirubin - (conjugated + unconjugated bilirubin)]. Newer technology like dry chemistry provide conjugated and unconjugated bilirubin fraction which is more specific and informative than direct and indirect bilirubin by conventional method in which only approximate values of conjugated bilirubin can be measured. In cholestasis conjugated bilirubin combines with albumin non enzymatically and form delta bilirubin in circulation and remains for long duration. The present review describes the usefulness of delta bilirubin in cholestasis, chronicity of liver diseases, in pediatric jaundice, transplant rejection and other jaundice condition.clinical correlation of delta bilirubin in all conditions have been summarized and their future perspectives are discussed.

A survey of the reactivity of in vitro diagnostic bilirubin reagents developed in Japan using artificially prepared bilirubin materials: A comparison of synthetic delta, unconjugated, and taurine-conjugated bilirubin.

In vitro diagnostic bilirubin reagents based on oxidation with bilirubin oxidase or vanadic acid for total and direct-reacting bilirubin are widely used in Japan; however, their reactivity to unconjugated and conjugated bilirubin and delta bilirubin has not been completely disclosed by manufacturers. We used artificially prepared bilirubin materials to investigate the reactivity with four in vitro diagnostic bilirubin reagents. Porcine unconjugated bilirubin solution, chemically synthesized ditaurobilirubin solution, and chemically synthesized delta bilirubin solution were used as surrogates of naturally occurring unconjugated bilirubin, conjugated bilirubin, and delta bilirubin, respectively. The total bilirubin and direct-reacting bilirubin concentrations were measured by three bilirubin oxidase methods and one vanadic acid method, and the observed concentrations were compared with those obtained by the diazo-based reference measurement procedure. The unconjugated bilirubin and delta bilirubin concentrations were similar when any of the four in vitro diagnostic bilirubin reagents were used during total bilirubin measurement. This was consistent with reference measurement procedure and exhibited a converged inter-method variation. Compared with reference measurement procedure, significantly low ditaurobilirubin concentrations were observed by the in vitro diagnostic bilirubin reagents despite the converged inter-method variation. In delta bilirubin measurement, some reagents reacted doubtfully with unconjugated bilirubin, while showed lower ditaurobilirubin concentrations than its corresponding total bilirubin concentration. Reactivity with delta bilirubin was different for each method including reference measurement procedure. Some reagents were developed to react less with delta bilirubin and others to strongly react with delta bilirubin. We revealed the reactivity of IVD-TB and IVD-DB reagents to artificially prepared bilirubin materials, and their consistency with reference measurement procedure. The delta bilirubin data results vary depending on the reagents used.

Clinical features of 9 cases of Brucella endocarditis

Objective: To explore the clinical characteristics of patients with Brucella endocarditis. Methods: The clinical data of 9 patients with Brucella endocarditis admitted to Beijing Ditan Hospital from October 2008 to August 2018 were retrospectively analyzed. Through the electronic medical record system of the hospital. Through assessing the electronic medical record system of the hospital, demographic data, main symptoms, vital signs, blood culture, Rose Bengal Plate Agglutination Test, echocardiography, electrocardiogram, chest imaging and other clinical data of included patients were inquired and recorded. Patients were followed up by telephone for medication, operation and outcome. Results: The 9 patients were all Han nationality, aged from 25 to 66 years, 7 out of 9 patients were male, and they came from Hebei, Shandong, Shanxi, Inner Mongolia and Beijing. Of the 9 patients, 5 were farmers, 2 were self-employed, 1 was a technician, and 1 was unemployed. Of the 9 cases, 8 had a history of close contact with cattle and sheep, and 5 had a history of eating beef and mutton. Rose-Bengal Plate Agglutination Test and blood culture were positive in all 9 patients. Aortic valve was involved in 7 out of 9 patients, mitral and tricuspid valve was involved in 1 patient, respectively, and aortic dissection occurred in 1 patient. Condition of 1 patient rapidly deteriorated after admission and finally died during hospitalization despite antibiotic therapy, the remaining patients received long-term antibiotic treatment. A total of 7 patients who underwent valve replacement were followed up. One patient died of cerebral hemorrhage 6 months after operation, and the remaining 6 patients recovered well after valve replacement. Heart failure occurred in all 9 patients, and pericardial effusion occurred in 8 patients. Electrocardiogram showed low voltage of the QRS complex in the limb in 3 cases and poor R-wave progression in V(1)-V(3) lead in 2 cases, and sinus tachycardia in 2 cases. One patient developed non-specific ST-T abnormalities. All patients had fever, 7 patients complained of weakness, and 6 patients complained of palpitations. Among the 9 patients, 7 cases had anemia, 7 patients had pneumonia, 6 had bilateral pleural effusion, 4 had thrombocytopenia. Creatinine was above normal in 4 patients, urine protein was positive in 3 patients, Delta Bilirubin was higher in 3 cases. Conclusions: Patients with Brucella endocarditis often suffer from heart failure and have severe complications. Adequate antibiotic therapy in combination with valve replacement is effective for the treatment of patients with Brucella endocarditis.

Characteristics of Bilirubin According to the Results of the Direct Antiglobulin Test and Its Impact in Hemolytic Disease of the Newborn.

Hyperbilirubinemia, which is a sign of hemolytic disease of the newborn (HDN), can irreversibly damage the central nervous system. To determine the etiology of HDN in affected patients and characterize the changing pattern of bilirubin using direct antiglobulin testing (DAT). We collected clinical data from newborns who underwent perinatal DAT and from their mothers, between August 2008 and July 2017. Among 303 neonates, 37 (12.2%) showed positive DAT results. The positive predictive values (PPVs) and negative predictive values (NPVs) based on DAT results were 75.7% and 28.9%, respectively, for starting phototherapy. Bilirubin levels increased more rapidly in the DAT-positive group, compared with the DAT-negative group. The initial bilirubin level differed significantly according to the etiology of hyperbilirubinemia. Further, neonates with anti-D showed higher delta bilirubin per day than neonates with other antibodies. Our results may help to determine the measurement period for bilirubin according to DAT results and etiology.

Delta Bilirubin as a Marker of Cholestasis

Delta Bilirubin as a Marker of Cholestasis

Comparison of Direct Bilirubin on Two Automated Systems: Influence of Delta Bilirubin

Comparison of Direct Bilirubin on Two Automated Systems: Influence of Delta Bilirubin

Biochemical Evaluation of Serum Bilirubin Fractions and Liver Function Tests in Sepsis Neonates with Hyperbilirubinemia

Background: Neonatal sepsis remains one of the leading cause of morbidity and mortality both among term and preterm infants. Sepsis and meningitis are responsible for most of these deaths. According to WHO estimates, there are about 5 million neonatal deaths a year. Jaundice and hepatic dysfunction frequently accompany a variety of bacterial infections. This study was aimed to evaluate bilirubin fractions and liver function tests in septic and non septic neonates with hyperbilirubinemia. Materials & Methods: A total of 41 neonates, their age ranged from (1-28 days), mean age sepsis cases 4.29±5.34 and mean age in non sepsis 2.27±4.44. The patients admitted to neonatology unit for the management of hyperbilirubinemia were included in this study. Out of 41, 20 babies having sepsis (17 were males & 3 were females) and 21 (15 were males & 6 were females) were non sepsis. All study subjects were studied for the serum bilirubin fractions and other liver function tests by using vitros dry chemistry analyzer. Results: In the present study, delta bilirubin (0.955 ±0.546) and conjugated bilirubin (1.17±2.10) levels are significantly increased in sepsis cases when compared to non sepsis controls. Conclusion: In conclusion, conjugated bilirubin and delta bilirubin were significantly increased in neonates suffering from sepsis with hyperbilirubinemia. By studying individual fractions of bilirubin, especially unconjugated bilirubin, conjugated bilirubin and delta bilirubin (not as direct and indirect bilirubin) will help in early diagnosis of sepsis and thus may help in better management of the sepsis neonates.

Concept of BuBc in neonatal jaundice – needs a change

Orthoclinical diagnostics Vitros microslide came with a method by which Bu and Bc can be estimated directly. The advantage of this microslide technology is that conjugated bilirubin (Bc) does not contain delta bilirubin and the fractions i.e BuBc are actually estimated. TBil estimation is also available based on traditional diazo method on the same instrument

Unexpected Presence of Delta Bilirubin in the Serum of a Patient with Type I Crigler-Najjar Syndrome Observed by a Kodak Ektachem 700N Analyzer

Unexpected Presence of Delta Bilirubin in the Serum of a Patient with Type I Crigler-Najjar Syndrome Observed by a Kodak Ektachem 700N Analyzer

PTU-091 Percutaneous transhepatic cholangiography (PTC); are we hitting the target?

IntroductionPercutaneous transhepatic cholangiography (PTC) is a procedure used to access the biliary tree for diagnostic purposes in obstructive jaundice, and to facilitate palliative stenting across biliary strictures. A previous study...

Infliximab monotherapy for severe alcoholic hepatitis and predictors of survival: An open label trial

Severe alcoholic hepatitis (AH) is associated with very high mortality. Tumor necrosis factor-alpha (TNF-alpha) contributes to the progression of AH and TNF-alpha antagonists like infliximab may help in ameliorating the severity and complications of AH. There is a scarcity of data regarding the safety and efficacy of infliximab monotherapy in the treatment of AH. We evaluated infliximab monotherapy in patients with severe AH. Patients with severe AH (Maddrey's score>32) received a single dose of infliximab 5 mg/kg IV. The primary endpoint was survival assessed at one and two months. The secondary endpoints were reduction of the Maddrey's DF and development of any bacterial infections. Predictors of survival were assessed at admission and at day 7. Nineteen patients were enrolled in the study and received infliximab. By the end of one month two patients died resulting in 1-month survival of 17/19 (89%). By the end of two months four additional patients died resulting in 2-month survival of 68%. At the end of one and two months, compared to baseline, there was significant improvement in median values of Maddrey's DF (p<0.05). Median serum TNF-alpha levels decreased from 45 (range 11-19,880) at baseline to 20 (range 4-8600) pg/mL at 4 weeks (p=0.001). CRP levels, MELD score, and absolute neutrophil count decreased significantly. Five patients (26%) developed infection: three of them had pneumonia, while two developed a flare of pulmonary tuberculosis. Three patients recovered with treatment but two patients (10%) died (one with pneumonia leading to sepsis and the other of disseminated tuberculosis). Absence of hepatic encephalopathy at admission significantly predicted survival. Among patients who survived only 1/13 (8%) had hepatic encephalopathy at admission while among patients who died 4/6 (67%) had hepatic encephalopathy (p=0.017). Lille score and delta bilirubin at day 7 (DBD7) (defined as [baseline serum bilirubin minus serum bilirubin at day 7] x 100/baseline serum bilirubin), also predicted 2-month mortality. The area under ROC curve of DBD7 values for predicting survival was 0.77 (95% CI 0.55-0.99). DBD7 of >7.5% best predicted survival in the patients (sensitivity 85%, specificity 67%, PPV 85%, NPV 67%, and overall accuracy 79%). In severe AH, single dose infliximab is associated with improvement in parameters of severity and survival. However, infection remains a concern. Hepatic encephalopathy at admission, Lille score and DBD7 predicted 2-month mortality. Large randomized controlled trials are needed before infliximab can be recommended for AH.

Conjugated Hyperbilirubinemia: Screening and Treatment in Older Infants and Children

1. Rula Harb, MD* 2. Daniel W. Thomas, MD† 1. *Children’s Hospital of Los Angeles, Los Angeles, Calif 2. †Editorial Board After completing this article, readers should be able to: 1. Describe the metabolism of bilirubin. 2. Evaluate a child of any age who has conjugated hyperbilirubinemia. 3. Recognize the signs and symptoms of Wilson disease. Jaundice refers to yellow discoloration of the skin, sclera, mucous membranes, and body fluids. It is a common problem that can be the presenting sign for many disorders. The challenge for the physician is to identify patients who need additional evaluation. The differential diagnosis for jaundice is age-specific; this review addresses the causative conditions in infants beyond the newborn period, older children, and adolescents. Jaundice is caused by elevated serum bilirubin concentrations. It is apparent in infants when the serum bilirubin value is greater than 4 to 5 mg/dL (68.4 to 85.5 mcmol/L) and in older children at values greater than 2 to 3 mg/dL (34.2 to 51.3 mmol/L). Serum total bilirubin is measured in the laboratory as the sum of two components: unconjugated (“indirect”) and conjugated (“direct”) fractions. The terms “direct” and conjugated hyperbilirubinemia often are used interchangeably. However, this usage is not always accurate because direct bilirubin may include both the conjugated fraction and bilirubin bound to albumin (delta bilirubin). Delta bilirubin is formed by covalent bonding between conjugated bilirubin in the serum and albumin; it is metabolized with albumin and has a similar half-life of 21 days. The presence of delta bilirubin often prolongs direct hyperbilirubinemia while results of the other liver tests are normalizing. Many hospitals continue to measure direct bilirubin by a method that includes both direct and delta bilirubin. Clinicians should consider asking for a breakdown of the direct bilirubin fraction if the jaundice is prolonged or presenting atypically. Conjugated hyperbilirubinemia is defined as a conjugated bilirubin concentration greater than 2 mg/dL (34.2 mmol/L) or more than 20% of total bilirubin. It …

Outcome from molecular adsorbent recycling system (MARS™) liver dialysis following drug‐induced liver failure

Fulminant liver failure from drug ingestion is associated with a high mortality, and the introduction of liver transplantation has improved the mortality significantly if done in a timely fashion. Recently, molecular adsorbent recycling system (MARS) liver dialysis has been introduced as a support for liver failure with varying results. We review our experience with drug-induced liver failure and the impact of MARS liver dialysis on the outcome, in a setting where cadaveric liver transplantation is rarely available. A total of 13 patients were treated, and 40 sessions of MARS liver dialysis were conducted in the intensive care unit. The majority of cases were because of herbal medicine toxicity. Total bilirubin, conjugated bilirubin, and delta bilirubin were significantly reduced, with no change in unconjugated bilirubin. All patients satisfied the criteria for urgent liver transplantation with an average Model End Stage Liver Disease (MELD) score of 35. Only one patient received a liver transplantation from a live donor (right lobe). Overall mortality was 85%. Median time-to-death from the start of MARS was 8 days. MARS liver dialysis in a setting without timely liver transplantation is associated with a poor outcome. It does, however, provide a window of time for consideration of living donors in the setting of limited cadaveric donors.

Rapid LC-TOFMS method for identification of binding sites of covalent acylglucuronide–albumin complexes

A method for rapid identification of binding sites of covalent adducts was developed using delta bilirubin as a model compound. Delta bilirubin, containing intact human serum albumin (HSA), was digested with trypsin and the peptide fragments were monitored at 436 nm, but no predominant peaks were detected indicating the instability of the digested peptides containing bilirubin-related compounds. Therefore, the high-performance liquid chromatography time-of-flight mass spectrometer (LC-TOFMS) data of digested fragments of delta bilirubin were compared with those of control digests of HSA, revealing a characteristic peptide in the digest mixture of delta bilirubin. This peptide was sequenced by high-performance liquid chromatography time-of-flight tandem mass spectrometry (LC-TOFMS/MS) and identified as LDELRDEGKASSAK (Leu182 to Lys195) with a modification of a 178 Da increase at Lys190. This indicated the Lys190 to be a predominant covalent binding site of BGs on HSA via the imine mechanism and the binding between the bilirubin moiety and the glucuronic acid moiety to be unstable to digestion with trypsin. The method of comparing LC-TOFMS data requires no specific detection such as fluorescence or radioactivity for every compound. This should accelerate the structure elucidation of covalent adducts and be helpful for studying the relationship between the structure of ligands and specific binding sites.

Accurate direct spectrophotometric bilirubin measurement combined with blood gas analysis

Background: A new total bilirubin (B T) method, based on multiple wavelength absorbance measurements, and an algorithm to calculate concentration, were evaluated for accuracy in specimens containing variable amounts of unconjugated bilirubin (B U), conjugated bilirubin (B C) and delta (protein-bound) bilirubin (B D). Methods: Quantitation of B U, B C, and B T (with calculation of B D) using a Vitros 250 analyzer served as the comparison method. Results: Analysis of neonatal specimens using a preliminary algorithm yielded good overall agreement with the Vitros B T method, but there was considerable variation in the agreement for individual specimens. When specimens from adults selected to yield a range of B C and B D levels were analyzed, the preliminary algorithm underestimated B T. Refinement of the method in the form of a finalized algorithm resulted in elimination of the negative bias seen with specimens with high B D and B C levels, and better agreement for individual neonatal specimens. Conclusions: This new method overcomes the limitations observed in earlier spectrophotometric methods, and provides accurate results in specimens containing a range of bilirubin forms.

Deproteinization of serum: another best approach to eliminate all forms of bilirubin interference on serum creatinine by the kinetic Jaffe reaction.

The negative interference of conjugated, unconjugated, and delta bilirubin on patient serum creatinine determined by the kinetic Jaffe reaction is the unresolved problem. We compared bilirubin interference on thirty patients' serum creatinine obtained from four analyzers, with and without deprotenization before the Jaffe reaction, to the Vitros dry enzymatic method. We found significant negative interference from bilirubin on serum creatinine in all samples directly applied to four wet chemical methods, except the one incorporated with serum blank rate. The negative interferences linearly related to bilirubin concentration. However, bilirubin did not interfere on serum creatinine obtained from all wet chemical methods incorporated with deproteinization process before the reaction. We conclude that deproteinized serum before the reaction is the best approach to eliminate all forms of bilirubin interference on serum creatinine determined by the kinetic Jaffe reaction.

The Development and Compensation of Biliary Cirrhosis in Interleukin-6-Deficient Mice

In an effort to understand the role of IL-6/gp130 signaling in chronic liver injury, IL-6 deficient (IL-6(-/-)) and wild-type control (IL-6(+/+)) mice were subjected to bile duct ligation (BDL) for 12 weeks. This maneuver causes chronic biomechanical stress and liver injury, fueling sustained biliary epithelial and hepatocyte proliferation. By 12 weeks after BDL, IL-6(-/-) mice develop significantly higher total serum bilirubin levels (23.2 +/- 2.3 versus 14.9 +/- 2.1 mg/dl, P < 0.0001; delta bilirubin subfraction 16.7 +/- 4.0% versus 9.2 +/- 1.8%; P < 0.002), and the majority (15/18) show "black" gallbladder bile, compared to IL-6(+/+) mice (5/16; P < 0.003). The IL-6(-/-) mice also cannot sustain the compensatory liver mass increase commonly seen with chronic obstructive cholangiopathy, because of less hepatocyte proliferation, despite a rate of hepatocyte apoptosis similar to that of IL-6(+/+) mice. Moreover, IL-6(-/-) mice show a more advanced stage of biliary fibrosis and a higher mortality rate than the IL-6(+/+) controls (51% versus 23%; P < 0.02). These phenotypic changes in the IL-6(-/-) mice are associated with decreased expression and phosphorylation of gp130 and the transcription factor STAT3, compared to IL-6(+/+) mice. Daily treatment with exogenous recombinant IL-6 for 3-6 weeks starting at 6 weeks after BDL significantly lowers the serum total bilirubin in both groups. In the IL-6(-/-) mice, exogenous IL-6 treatment also increases the level of gp130 protein expression and completely reverses the loss of liver mass by increasing the hepatocyte proliferation. In conclusion, IL-6 appears to contribute to biliary tree integrity and maintenance of hepatocyte mass during chronic injury.

Evaluation of delta bilirubin in the follow-up of hepatic transplantation

Evaluation of delta bilirubin in the follow-up of hepatic transplantation