EP1248951A2 - Diagnostiches verfahren zur verwendung von hämaglobin, hämatokrit oder rotblutzell-anzahl - Google Patents

Diagnostiches verfahren zur verwendung von hämaglobin, hämatokrit oder rotblutzell-anzahl

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Publication number
EP1248951A2
EP1248951A2 EP01901248A EP01901248A EP1248951A2 EP 1248951 A2 EP1248951 A2 EP 1248951A2 EP 01901248 A EP01901248 A EP 01901248A EP 01901248 A EP01901248 A EP 01901248A EP 1248951 A2 EP1248951 A2 EP 1248951A2
Authority
EP
European Patent Office
Prior art keywords
animal
levels
disease
haematocrit
haemoglobin
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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Application number
EP01901248A
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English (en)
French (fr)
Inventor
Toby Grahame Knowles
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
University of Bristol
Original Assignee
University of Bristol
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by University of Bristol filed Critical University of Bristol
Publication of EP1248951A2 publication Critical patent/EP1248951A2/de
Withdrawn legal-status Critical Current

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/72—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving blood pigments, e.g. haemoglobin, bilirubin or other porphyrins; involving occult blood
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/5005—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
    • G01N33/5091—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing the pathological state of an organism

Definitions

  • the present invention relates to a method of predicting the growth potential of an animal and/or the ability of an animal to resist disease, particularly transmissible diseases.
  • the methodology is simple to apply, results are almost instantaneous and the procedure is non-traumatic.
  • the ability to predict disease or growth rate in young stock is of use in tailoring a system of husbandry to best economic advantage.
  • the inventors have carried out studies of the reference ranges for blood biochemical and haematological variables in young animals and have defined a surprising correlation between the blood biochemical and haematological measurements obtained and the future growth potential, as relative percentage live weight gain over time, and the ability of the animal to resist disease, in terms of a normal (not raised) white blood cell count. (Knowles T. G. et al (2000), The Veterinary Record, 147, 593-598). Specifically, measurement of red blood cell count (RBC), haematocrit and blood haemoglobin have been shown by the inventors to be useful in such a predictive role at an early stage of an animal's life: The physical measurement of these blood parameters may be performed by any known procedure.
  • the invention predicts whether an animal is more susceptible to transmissible diseases, i.e diseases which result in an immune response (as evidenced by a raised white blood cell count and/or neutrophil count).
  • transmissible diseases i.e diseases which result in an immune response (as evidenced by a raised white blood cell count and/or neutrophil count).
  • Such diseases are generally a. result of infection by bacteria, mycobacterium, viruses or other biological agents and can take the form of gastrointestinal disease, respiratory disease, infection of any part of the body by a biological agent.
  • the invention can be used at birth or within the first week of life to predict disease susceptibility in later life.
  • a method of predicting the ability of an animal to resist disease comprising:
  • the disease is a transmissible disease for example respiratory disease, enteritis, pneumonias, corona virus, joint-ill, navel-ill, staphlococcal infection, streptococcal infection, or pasteurellosis infection.
  • a transmissible disease for example respiratory disease, enteritis, pneumonias, corona virus, joint-ill, navel-ill, staphlococcal infection, streptococcal infection, or pasteurellosis infection.
  • Other variables include serum albumin levels, alkaline phosphatase levels, ⁇ -hydroxybutyrate levels, plasma cortisol levels, serum creatine kinase levels, creatinine levels, iron levels, plasma fibrihogen levels, serum ⁇ glutamate levels, plasma glucose levels, haptoglobin levels, serum non-esterified fatty acid levels, total protein levels, transferrin levels, triglyceride levels, urea, ⁇ globulin, basophil levels, eosinophil levels, packed cell volume levels, haemoglobin levels, lymphocyte levels, mean cell haemoglobin levels, mean cell haemoglobin concentration, mean cell volume levels, monocyte levels, band neutrophil levels, neutrophil levels, platelet counts, and white blood cell count.
  • red blood cell count (RBC)
  • the measured RBC in counts of 10 12 /1
  • the predicted disease susceptibility of an animal using the formula:
  • a score of less than 0 indicates that the animal has a higher than normal predicted disease susceptibility.
  • the measured haematocrit level (as a percentage) can be correlated with the predicted disease susceptibility of an animal using the formula:
  • the measured haemoglobin levels (in units of g/dl) can be correlated with the predicted disease susceptibility of an animal using the formula:
  • a score of less than 0 indicates that the animal has a higher than normal predicted disease susceptibility.
  • the inventors have found that the higher the predicted level of disease susceptibility, the lower the growth potential of the individual animal.
  • a diagnostic kit for use in a method of determining the ability of an animal to resist disease comprising means to measure the level of one or more variables present in a blood sample where the variables are selected from haematocrit levels, red blood cell count or haemoglobin levels.
  • Other variables include serum albumin levels, alkaline phosphatase levels, ⁇ -hydroxybutyrate levels, plasma cortisol levels, serum creatine kinase levels, creatinine levels, iron levels, plasma fibrinogen levels, serum ⁇ glutamate levels, plasma glucose levels, haptoglobin levels, serum non-esterified fatty acid levels, total protein levels, transferrin levels, triglyceride levels, urea, ⁇ globulin, basophil levels, eosinophil levels, packed cell volume levels, lymphocyte levels, mean cell haemoglobin levels, mean cell haemoglobin concentration, mean cell volume levels, monocyte levels, band neutrophil levels, neutrophil levels, platelet counts, and white blood cells count.
  • a method of predicting the growth potential of an individual animal comprising:
  • a taking a blood sample from the animal; and b. measuring the level of one or more variables selected from haematocrit levels, red blood cell count or haemoglobin levels; and c. using the level determined in (b) to predict the growth potential of the animal.
  • the animal may be from a domesticated species.
  • the animal is a calf.
  • the animal may be less than one year old.
  • the blood sample is taken from birth to within the first three days after birth.
  • the animal may be a human.
  • the animal is less than one year old.
  • the blood sample is taken from birth to within the first three days after birth.
  • a fourth aspect of the invention provides a diagnostic kit for use in a method of determining the growth potential of an animal comprising means to measure at least one of haematocrit, red blood cell count and/or haemoglobin in a blood sample.
  • Figures 1 (A) to 1 (F) are graphs showing the changes in levels of (A) albumin, (B) ALP,
  • Figures 2 (A) to 2 (F) are graphs showing the changes in (A) Fe, (B) fibrinogen, (C) GGT,
  • Figures 3 (A) to 3 (F) are graphs showing the changes in (A) NEFA, (B) total protein, (C) transferin, (D) triglycerides, (E) urea and (F) ZST in calves from birth to 83 days; and
  • Figures 4 (A) to 4 (F) are graphs showing the changes in (A) haematocrit, (B) haemoglobin, (C) lymphocytes, (D) mean cell HB, (E) mean cell HB CN and (F) mean cell volume in calves from birth to 83 days; and
  • Figures 5 (A) to 5 (E) are graphs showing the changes in (A) monocytes, (B) neutrophils, (C) platelets, (D) RBC and (E) white cells in calves from birth to 83 days.
  • calves Fourteen calves were blood sampled from birth to 83 days of age. A blood sample was taken within three hours of birth and at 1, 3, 6, 9, 13, 20, 27, 41, 55 and 83 days of age. On each occasion a measurement of girth, from just behind the forelimbs, was also taken. A colostrum (first milk) sample was obtained from the dam as soon as possible post partum, for determination of specific gravity. The specific gravity of the colostrum is related to the amount of immunoglobulin present in the milk. The measurement of ZST in the blood was used to ensure that transfer of the immunogloblins from the milk to the calf had indeed taken place. The calves were kept and sampled on the farms on which they were born, with no alteration of the usual husbandry practices of the farm.
  • a 10ml blood sample was obtained by jugular venipuncture using an uncoated 10ml monovette.
  • the sample was then split into five parts; approximately 2ml was placed in each of two 2ml EDTA vacutainers (purple top), a 2ml Lith/Hep vacutainer (green top), a 2ml OxF vacutainer (grey top) and a 2ml uncoated vacutainer (red top).
  • Different vacutainer coatings were required as the different assays required that the blood be preserved in different ways. The aim was to mimic, as closely as possible, the treatment that samples would be subjected to when obtained by veterinarians in the field.
  • the tubes for biochemical analysis were taken to the Langford Veterinary Investigation Centre analysis by the Veterinary Laboratory Agency (VLA). Where samples were obtained at the weekend they were stored at 4°C before delivery to the VLA on the first working day.
  • the tube for haematological analysis was analysed by the Langford haematology laboratory. Where samples were obtained at the weekend, two whole blood slides were prepared immediately and the remainder of the sample was stored at 4°C before analysis on the first working day. The remaining Lith/Hep tube was centrifuged within 60 minutes of collection and the plasma recovered and stored at -20°C for later analysis of cortisol levels.
  • the samples delivered to the VLA were analysed for levels of albumin, alkaline phosphatase (ALP), ⁇ -hydroxybutyrate (BHB), creatine kinase (CK), creatinine, iron (Fe), fibrinogen, ⁇ -glutamyltransferase (GGT), glucose, haptoglobin, non-esterified fatty acid (NEFA), total protein, transferrin, triglycerides, urea and ⁇ globulin (as measured by the zinc sulphate turbidity test, ZST).
  • ALP alkaline phosphatase
  • BHB ⁇ -hydroxybutyrate
  • CK creatine kinase
  • CK creatinine
  • Fe iron
  • GTT ⁇ -glutamyltransferase
  • NEFA non-esterified fatty acid
  • ZST zinc sulphate turbidity test
  • Haematological analyses included eight variables: haematocrit, haemoglobin level, mean cell haemoglobin (mean cell HB), mean cell haemoglobin concentration (mean cell HB CN), mean cell volume, numbers of platelets, red blood cells (RBC), and white cells. Samples were run through an automated impedance cell counter (Baker 9000) to obtain the 'eight parameter, complete blood count'. Blood smear slides were stained with Leishmans' stain (1ml stain for 3 minutes, 2ml 6.8 pH buffered, distilled water for 12 minutes, rinsed in buffer and then allowed to air dry).
  • the manual leukocyte differential count was performed by counting and identifying 100 cells using the 'battlement counting technique' (Jain 1986) to produce counts of band neutrophils (bands), basophils, eosinophils, lymphocytes, monocytes and neutrophils. Bands, basophils and eosinophils are mentioned further later as the majority of counts for these variables were zero.
  • HWC high white cell count
  • LWC low white cell count
  • the mean counts (x 10 "9 ) are given and the number of animals upon which the mean is based is given below.
  • the figures in brackets show the number of animals which had at least one positive count throughout the survey.
  • HWC and LWC groups differ in terms of other variables; especially so because the differences between the groups in some of the variables are apparent at birth and these predict the immune response to disease for example by the raised WBC count.
  • Table 4 shows the normal ranges of the biochemical variables, for cattle, issued by the VLA and the normal ranges of the haematological variables given in Jain, N. C. supra (1984) and Radostits, O. M., et al (1994) Veterinary Medicine: A Textbook of the Diseases of Cattle, Sheep, Pigs, Goats and Horses. Bailliere Tindall. ISBN: 070201592X. These can be compared with the results obtained from this study as an indicator of where values in healthy calves are likely to deviate from the published normal values. Where possible the values for the normal range are indicated in Figures 1 to 5 by a horizontal line. Where the normal value is/are off scale an arrow head indicates the direction of the value(s). There are no normal ranges shown on the graphs of cortisol, girth or transferrin.
  • Triglyceride levels in the calves were always within the reference values for cattle. Levels peaked at birth at 0.4 to 0.5 mmol/1 but after day 3 remained roughly in the range 0.2 to 0.3 mmol/1.
  • Lymphocytes Mean cell HB, Mean cell HB CN and monocytes The values of all these variables were within the published reference ranges for cattle but all showed marked patterns of change from birth to 83 days.
  • the error bars shown on the Figures can be used to give the 'normal reference range' at a particular time, as the standard error of the mean is equal to the standard deviation divided by the square root of the number animals in a sample. As mentioned earlier, normal reference ranges can be based on plus or minus two standard deviations about the mean.
  • Table 5 Estimated logistic regression equations for assigning group membership using the blood variables RBC, haematocrit and haemoglobin, individually. The cut-off point is 0. Thus a score of less than 0 indicates membership of the disease susceptible group.
  • Tables 9 to 14 show the raw data of measurements of RBC, haematocrit, haemoglobin, girth (a measurement of growth), white cell count (WCC) and neutrophil count (NC) (measurements of immune reaction), respectively, from calves ⁇ Bos torus) A to N, at birth, and on days 1, 3, 6, 9, 13, 20, 27, 41, 55 and 83 of life.
  • a measurement of the red blood cell count (RBC) of an animal within the first few days of birth can be used to predict growth potential and disease resistance. Animals with counts below 10.38 x 10 ⁇ /litre are classified as having better growth potential and better disease resistance.
  • the method may be adapted for more accurate use on specific animal populations by means of sampling the population of interest and adjusting the cut-off point for RBC accordingly.
  • Other measurements of blood such as, haematocrit (also known as packed cell volume (PCV)) and blood haemoglobin which are highly correlated with red blood cell count may be used in a similar predictive role.
  • the cut-off points for haematocrit and haemoglobin are 48.6 per cent and 14.5 g/dl, respectively. Measurements below these values predict better growth potential and better disease resistance.
  • the invention may be implemented using any of the commercially available methods for measuring these parameters in blood.
  • Table 6 Results of the logistic regression using either RBC, haematocrit or haemoglobin as predictors of group membership. The statistic showing the overall goodness of fit is shown together with the parameter estimates of the model and the percentage of the animals correctly classified into the two groups.
  • Table 7 The multivariate results of a repeated measures analysis of variance showing a trend for differences in growth rate between the high and low white cell count groups over time (see Figure 2D).

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Hematology (AREA)
  • Immunology (AREA)
  • Molecular Biology (AREA)
  • Biomedical Technology (AREA)
  • Chemical & Material Sciences (AREA)
  • Urology & Nephrology (AREA)
  • Biotechnology (AREA)
  • Biochemistry (AREA)
  • Cell Biology (AREA)
  • Food Science & Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Microbiology (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Physiology (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
EP01901248A 2000-01-17 2001-01-17 Diagnostiches verfahren zur verwendung von hämaglobin, hämatokrit oder rotblutzell-anzahl Withdrawn EP1248951A2 (de)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
GB0001058 2000-01-17
GB0001058A GB0001058D0 (en) 2000-01-17 2000-01-17 Diagnostic method
PCT/GB2001/000173 WO2001053830A2 (en) 2000-01-17 2001-01-17 Diagnostic method involving haemoglobin, haematocrit or reg blood cell count

Publications (1)

Publication Number Publication Date
EP1248951A2 true EP1248951A2 (de) 2002-10-16

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EP01901248A Withdrawn EP1248951A2 (de) 2000-01-17 2001-01-17 Diagnostiches verfahren zur verwendung von hämaglobin, hämatokrit oder rotblutzell-anzahl

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EP (1) EP1248951A2 (de)
AU (1) AU771538B2 (de)
GB (1) GB0001058D0 (de)
WO (1) WO2001053830A2 (de)

Family Cites Families (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0115459A3 (de) * 1983-01-26 1986-11-20 University Of Medicine And Dentistry Of New Jersey Haematologische Voraussage von Sepsis und anderen erkrankten Zuständen
RU1772746C (ru) * 1990-07-10 1992-10-30 Смоленский государственный медицинский институт Способ исследовани динамики отека головного мозга
US5506145A (en) * 1994-12-02 1996-04-09 Bull; Brian S. Determination of an individual's inflammation index from whole blood fibrinogen and hematocrit or hemoglobin measurements
RU2123697C1 (ru) * 1996-08-05 1998-12-20 Алтайский государственный медицинский университет Способ прогнозирования исходов тяжелых воспалительных заболеваний челюстно-лицевой области

Non-Patent Citations (1)

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Title
See references of WO0153830A3 *

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AU771538B2 (en) 2004-03-25
AU2690801A (en) 2001-07-31
WO2001053830A2 (en) 2001-07-26
GB0001058D0 (en) 2000-03-08
WO2001053830A3 (en) 2001-12-27

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