Ethical approval: The experiments on animals were approved by Animal Welfare and Experimentation Ethics Committee (AWEEC), Bangladesh Agricultural University, Mymensingh-2202.
Animal sampling: A total of 20 Black Bengal goats of different ages were collected randomly from local market, Mymensingh sadar, Bangladesh. All of the Black Bengal goats were considered clinically healthy animals at the time of sampling and all of them were fed on the same diet.
Blood sampling: Blood samples were collected from the jugular vein into two tubes (Guangzhou Improve Medical, China), in which one contained anticoagulants (ethylene di-amine tetra acetic acid) for blood hematology and the other tube contains no anticoagulants (Becton Dickinson, Franklin Lakes, USA) for the biochemical analysis. All of the samples were transferred into the laboratory as quick as possible in ice.
Blood hematology: Blood hematology prepared by using a special blood lysing buffer approved for goat hematology (Concentrated Lysing Reagent, SEAC, Florence, Italy). All the samples were analyzed within 45 min after collection by using CELL-DYN 3700 analyzer for the total red blood cells count (RBC), total and differential white blood cells count (WBC), hemoglobin (HGB), hematocrit (HCT), mean corpuscular volume (MCV), mean corpuscular HGB (MCH), MCH concentration (MCHC), red cell distribution width, and mean platelet volume.
Serum biochemistry: The samples were left to clot and then centrifuged at 3000 rpm for 15 min, and the serum is collected. Serum was kept frozen at −20°C until it was used for the biochemical analysis. Serum biochemistry was carried out by using Vet Scan VS2 analyzer (ABAXIS, USA) for the following parameters: Albumin (ALB), alanine aminotransferase (ALT), alkaline phosphatase (ALP), gamma glutamyltransferase (GGT), and blood urea nitrogen (BUN). Serum Immunoglobulins were analyzed by ELISA method, using auto AnalizerMatchine (BN Prospect, Siemens Healthcare) for humoral immune responses at different ages in Black Bengal goat.
Statistical analysis: For the statistical analysis, GraphPad Prism 7/ GLM of SAS (2010) version 9.3.software is used to calculate the minimum and maximum values to determine the range, mean, standard deviation of the mean also Shapiro–Wilk normality test was used to evaluate the normal distribution of the values. All the resulting data for the blood metabolites was checked for normality using the PROC UNIVARIATE of SAS (2010). A repeated measures analysis of SAS (2010) was used to assess the effect of time on blood metabolites and haematological parameters.