IntroductionrnReference ranges are values for analytes that aid physicians inrnaccurate diagnosis proper treatment and follow-up of patients. Theyrnrepresent a defined group of individuals. When doing a follow uprnon patients a clinician often uses a subject-based reference value torndetermine the progress made in the management of a pathologicalrndisorder 1. To establish whether a patient has a certain pathologicalrndisorder however group-base reference range is used in therninterpretation of laboratory report. Correct interpretation of the resultsrnfrom these analyte presupposes that the clinician and the laboratoryrnmedicine physician have good reference information 2. This grouprnshould be as similar as possible to the patients under investigation.rnThe reference population is recruited from the individuals who fulfilrna defined inclusion and exclusion criteria as well as a defined partitionrncriterion. The samples from the reference population are then sortedrnbased on the results of inclusion and exclusion criteria. The resultingrngroup of individuals is regarded as a reference population and thernresults were evaluated statistically to establish the reference values. Thernupper and lower limit of reference value measurements is dependent onrnage sex genetics diet altitude and the methodology used. In additionrnreference values produced by reagent manufacturers are determinedrnfrom analysis of blood samples of a few health workers who do notrnrepresent the general population.rnFrequently adult reference intervals are not appropriate forrnpediatric patients. To assist physicians in treating their pediatricrnpatients in Kenya paediatric reference values should be used. Currentlyrnthere are no established population based Kenyan reference limits forrnpaediatric patients. The reference limits in use in Kenya are eitherrnborrowed from the textbooks and articles or insert literature from thernkit manufacturers 1. It is therefore recommended that each clinicalrnchemistry laboratory establish its own paediatric reference range forrnbiochemical parameters. The objective for this study was to determinernreference ranges for some biochemical parameters for paediatricrnpopulation from Taita Taveta County Kenya 3.
The objective for this study was to determinernreference ranges for some biochemical parameters for paediatricrnpopulation from Taita Taveta County Kenya 3.rnMaterials and MethodsrnArea of studyrnThis was done at Moi District Hospital Voi Taita Taveta CountyrnKenya. Voi is situated in a sisal farming area in the Coastal region ofrnKenya.
Materials and MethodsrnArea of studyrnThis was done at Moi District Hospital Voi Taita Taveta CountyrnKenya. Voi is situated in a sisal farming area in the Coastal region ofrnKenya.rnParticipantsrnThe study group consisted of normal and healthy infants andrnchildren aged between 1 to 17 years and had stayed in Taita TavetarnCounty for not less than six months. Infants and children whosernparents and/or guardians consented to participate in the study. Therncriterion for selection was based on the inclusion criteria.rnEthical considerationrnKenyatta University Ethical Review Committee KUERC andrnMinistry of Health Wundanyi District Kenya granted the permissionrnto carry out the study. Inclusion and exclusion criteriarnSamples for inclusion in the study were to be from normal malesrnand females aged between 1 and 17 years who were residing in TaitarnTaveta County Kenya and had stayed for more than six months.rnThe exclusion criteria were subjected to all samples that were positivernfor Syphilis HIV antibody and Hepatitis B antigen. In addition allrnsamples collected from pregnant adolescent females were excluded.rnFinally adolescents whose parents had not consented to participatingrnin this study.rnImmuno-chromatographic reagent strip Determine HIV-1/2rnTokyo Japan was used for screening of HIV 1 and 2.50 L of thernsample was applied to the sample pad. After 1 minute chase buffer wasrnapplied to the sample pad and the test results read within 15 minutes.rnPositive results were indicated by the appearance of two red bars eachrnon the control window and on the patient window. Negative resultsrnwere indicated by the appearance of only one red bar at the controlrnwindow.rnThe HbsAg one step Test Strip HBsAg Beijing China wasrnused for the screening of HbsAg. This was a qualitative lateral flowrnimmunoassay test. The test strip was immersed in a tube containingrnthe serum for screening for 10 to 15 minutes. It was then removed andrnplaced on a non-absorbent flat surface and the results read within 15rnminutes. Positive result was indicated by the appearance of two distinctrnred bars one on the control region and the other on the test region.rnNegative results were indicated by the appearance of only one red barrnat the control window.rnThe syphilis ultra-rapid test which is a qualitative membrane striprnbased immunoassay Treponema pallidum Strip Beijing China wasrnused for the screening of Treponema pallidum which is the causativernagent of the venereal disease syphilis. 50 L of the serum sample wasrnplaced on the sample pad followed by 1 drop of buffer. The result wasrnread after 10 minutes. Positive result was indicated by the appearancernof two red lines one on the control region and the other on the testrnregion. Negative result was indicated by the appearance of one red linernon the control region.rnSpecimen collectionrnSamples from healthy infants and children were collected betweenrnthe months of July and November 2012. Five millilitres of blood wasrndrawn with the aid of a syringe and needle for children above five yearsrnand a scarp vein and syringe for children below five years. This was thenrntransferred to plain vacutainer tubes labelled with the subjects namernand study number. The blood samples were placed in ice-cool box readyrnfor transportation to the laboratory for analysis. In the laboratory thernblood samples were subjected to both inclusion and exclusion criteria.rnSpecimen transportation processing and storagernSpecimens collected were transported processed and stored accordingrnto the standard cold chain procedures as done by Waithaka et al. 1
Laboratory analysisrnTen renal function tests done included: Uric acid UA creatinernkinase CK urea BUN amylase AMYL creatinine CREATrnand electrolytes were determined on the serum processed fromrnblood samples. Standard procedures prepared by the Moi DistrictrnHospital Voi were used for this study. All tests were done using CobasrnIntegrarn 400 plus automatic Chemistry Analyzer Roche DiagnosticsrnMannheim Germany.rnCalibration of testsrnCalibrator for automated systems C.f.a.s was used. The systemrnperformed calibrations automatically.rnQuality assurance QA/Quality control QCrnTo ensure accuracy and precision of the test results all pre-analyticalrnanalytical and post analytical precautions were taken into consideration.rnInternal QC materials Precinorm and Precipath were procuredrnfrom Roche diagnostics and run according to the instructions of thernmanufacturer. Similarly the American Proficiency Institute API externalrnQC material was used for monitoring performance and participation atrnthe Moi District Hospital Laboratory and run twice within the periodrnof analysis. This was performed according to the instructions of thernmanufacturers and QC protocols. Quality control was done automaticallyrnas defined in the test specification for every analysis 4.rnData management and statistical analysisrnThe data was entered into a spread sheet and cleaned. Box plot wasrnused for identification of outliers 5. The data was then uploaded to SPSSrnv.20 for determination of the quartiles and interquartile range. Outliersrnwere determined using the procedure adopted by Waithaka et al. 1.rnA test of normality was also done 6. Reference values and mediansrnwere calculated at 95 confidence interval. Comparison for variationsrnbetween the genders was done using Mann Whitney test. Significantrndifferences were determined at a value of p 0.05. Comparison forrndifferent ages on the basis of gender was determined using One WayrnAnova followed by Dennetts test for multiple comparisons 7.
ResultsrnEstablishment of reference ranges for infants and children ofrnTaita Taveta County KenyarnAll the renal function tests RFTs comprising Urea creatininernPotassium and Chloride were similar while sodium showed significantrnvariation on the basis of sex p 0.05. There was also variation onrnthe basis of gender for uric acid while calcium and Phosphorous forrnboth genders were similar. The enzyme Amylase showed no significantrnsex differences p0.05 whilst Creatinine kinase showed significantrnsex difference p0.05 Table 1. During the entire analytical periodrneveryday control value result and the standard deviation SD from therncontrol target value were noted. All the daily QC runs were within rn2SD from the target values.rnReference values for healthy infants and children in differentrnage groups for Taita Taveta County KenyarnIn children aged between 6-10 years male children had significantlyrnhigher levels of Urea BUN and Creatinine kinase than their femalerncounter parts. In children aged between 11-15 years male children hadrnsignificantly lower levels of Phosphorous and significantly higher levelsrnof Urea and Creatinine kinase than their female counter parts. Malerninfants and children had: a significantly decreased level of PotassiumrnAmylase and Sodium in children aged more than 15 years comparedrnto children aged between 1-5 years old Table 2.rnFemale infants and children had: a significantly decreased level ofrnPhosphorous and a significantly increased level of uric acid in children aged more than 15 years compared to children aged 1-5 years old arnsignificantly decreased level of Phosphorous in children aged more thanrn15 years compared to children aged between 6-10 years a significantlyrndecreased level of Phosphorous and a significantly increased level ofrnUric acid in children aged more than 15 years compared to childrenrnaged between 11-15 years Table 2. The biochemical parameters:rnCalcium Chloride Creatinine Kinase and Urea are not affected byrnthe age of infants and children Table 2.rnThe results for significance difference in females for Uric acidrnbetween the four groups 1-5 years 6-10 years 11-15 years and over 16rnyears Table 3 while there was no significance in different age groups inrnPotassium Calcium Amylase Urea Sodium Chloride and CreatinernKinaseat p0.05. For the Phosphorous Potassium Calcium AmylasernUrea Sodium Chloride Uric acid Creatinine Kinase and Creatininernshowed no significance difference between age groups for the malernpopulation of ages 1-17 years at p0.05. In there was no significancerndifferences in both male and female of all age groups for all the analytesrnPotassium Calcium Amylase Urea Sodium Chloride GGT UricrnAcid Creatinine Kinase and Creatinine at p0.05 Table 4. Male and females of age 6-10 showed significance deference inrntheir references ranges for Uric acid and Creatinine Kinase whilernKinase Calcium Amylase Urea Sodium Chloride and Creatininernat p0.05 Tables 3 and 4. Significance difference was shown inrnUrea Potassium and Creatine kinase for ages 11-15 years for bothrnmale and females while no significance was noted in PotassiumrnCalciumAmylase Sodium Chloride Uric acid and Creatinine Tablesrn3 and 4 at p0.05. For ages above 16 years at P0.05 there was nornsignificance was noted in Potassium Calcium AmylaseL SodiumrnChloride Urea Creatinine Kinase Uric acid and Creatinine. Forrncomparison with references in literature there was no establishedrnsignificant differences found in the same age bracket 1-5 years 6-10rnyears 11-15 years and 16-17 years as those in the study. During thernentire study period everyday control value result and the standardrndeviation SD from the control target value were noted. All the dailyrnQC runs were within 2SD from the target values Figures 1-8.rnDiscussionrnThis study provides the first established clinical chemistry reference values for children aged 1-17 years for both males and females inrnTaita Taveta County Kenya derived from healthy individuals. Out ofrn577 participants recruited only 553 were involved in the study. Thernnumber of males 276 was almost equal to that of females 277rneach group exceeded the minimum of 120 participants per subgrouprnfor nonparametric estimates required for 95 reference intervalrndetermination as recommended by CLSI 7. Both the internal andrnexternal controls were used for quality control of the study besidesrnusing procedures set at the facility 4. in these values for different genders for these parameters. Other studiesrnreport similar findings for adult black populations in east Africa andrnUSA 8-12. It is shown that males have higher values for Creatininernkinase compared to females contributed by a higher mass of musclesrnand bone mass 1314. As a child grows the muscle mass increasesrnbut mechanical shock to muscle decreases and therefore explaining therndecrease in Creatinine Kinase as the children grow from 1-17 years duernto reduction in activity.rnThe variations in values for Sodium and Uric acid in the differentrnsexes may be associated with the different responses to dietary salts.rnThis may be explained by the variations in the sex hormone and geneticrnfactors which vary in the different sexes. Other similar findings in adultrnpopulations have been determined in Rwanda 9. Variation in uricrnacid have also been done 81011. This is suggested to be as a resultrnof differences in sex hormones and body mass. The observed variationsrnin the biochemical analytes in different ages and sexes may be anrnindication that some of the parameters are age dependent. The increasernin serum reference range for Uric acid in males with progression ofrnage could be as a result of the increase in weight with advancing age asrnsuggested by Kuzuya et al. 15. The decrease in serum reference rangernfor Amylase in females with progression of age could be partly due torna decrease in pancreatic function and integrity with age and geneticsrnThis is in agreement with other study findings 16. The significancernincrease in Phosphorous in early stages in children life is attributed tornbone growth but this reduces as they approach adulthood as the bonerngrowth has reduced. This can also be affected by diet. Low vitamin Drnand high calcium have some effect on Phosphorous.rnBoth kidney evaluation parameters vary with those in manufacturesrnreference ranges 8 this could be attributed to the fact that thernorgans are not fully developed. Amylase was extremely high thatrnmanufacturers reference ranges 8 but as they approach the age of 17rnyears they concur with manufacturers ranges. From the research it wasrnnoted that there was variation in as per age sex in all categories thisrncould be contributed to diet environmental factors 17 and analyticalrnmethods as indicated by E er e 10.rnDifferent lifestyles and genetic composition of different populationsrncould also explain the differences among the different genders andrnwithin the same genders as females and males as shown in Tables 3 andrn4 respectively. These differences have also been reported from otherrncountries 101118.rnGenerally physiological functions have been shown to vary withrnpopulation due to differences in diet genetics physical environmentalrnand socio-economic conditions 1920.rnThe reference values for most analytes determined in this studyrnvary from the same population. This indicates that there is need torndetermine gender and age established values which will be applicablernto specific populations for different geographical regions with differentrndiets and minerals salts present in the soils rather than using referencernvalues determined for all population from different geographicalrnregions and applied for all people 20.
ConclusionrnThe study has enabled the renal cardiac and pancreatic referencernranges for Taita Taveta Kenya which is independently from those thatrnare quoted by reagents manufacturers in addition to those quoted inrnmedical books. There was evidence that some of the parameters varyrnwith those in literature. This has been supported by similar studiesrndone in other counties and with different population all over the world.