The role of technology in hemoglobin testing has led to the creation of innovative devices and techniques for quantifying hemoglobin concentration in patients.
Using an automated hemoglobin analyzer, for instance, has enabled fast, accurate, and reliable results in hematology. A hemoglobin test may rely on different techniques, including reagent-based and “reagent-less” methods, or a variety of non-invasive procedures.
Hemoglobincyanide (HiCN) Method
The HiCN method of hemoglobin measurement can proceed using the principle of hemoglobin conversion to cyanmethemoglobin by adding ferricyanide and potassium cyanide.
The advantage of this type of hemoglobin test is the availability of an internationally accepted reference standard calibrator [1]. The HiCN method is still routinely used in clinics, especially in resource-poor countries; however, its time-consuming and cyanide-dependent protocol makes it more relevant as a reference method for POC hemoglobin devices and analyzer calibration.
Vanzetti’s Azide Methemoglobin
Hemoglobin conversion via potassium ferricyanide to the colored, stable azide methemoglobin form with an almost identical absorbance spectrum as HiCN, representing the key characteristic of Vanzetti’s azide methemoglobin method [2].
The HiCN reference method uses a similar reagent, except that sodium azide is substituted for potassium cyanide. This hemoglobin measurement method shows high specificity and sensitivity [3].
Point-of-care (POC) hemoglobin measurement devices, such as the HemoCue® 201 and EKF Hemo Control, operate using a modified version of Vanzetti’s method. First, blood is pulled into a dry reagent cuvette by capillary action. Second, the reagent destroys the red blood cell walls. Free hemoglobin is then oxidized to methemoglobin and finally converted to azide methemoglobin. The reagent’s susceptibility to humidity represents a challenge associated with this method.
“Reagent-Less” Methods
Due to the limitations of reagent-based cuvettes, POC devices have been developed with “reagent-less” cuvettes. HemoCue® 301 was the first POC device to possess such a feature. This device quantifies the absorbance of oxygenated and deoxygenated hemoglobin, while turbidity is measured and compensated for at 880 nm.
Currently an EKF Diagnostics company, DiaSpect represents a leader in new technology that measures hemoglobin without a reagent based on broad-spectrum photometry. The DiaSpect technology flashes a white LED light through a sample to an optical sensor component.
This sensor element identifies the blood’s absorbance at a broad wavelength range, providing insight into the overall absorbance spectrum, resulting in higher specificity and lower sensitivity to interference. “Reagent-less” Diaspect cuvettes feature extensive durability, with a total shelf-life of 2.5 years. Just about one second of measurement time is another benefit of this type of device.
Non-Invasive Methods
With new technologies to detect hemoglobin’s spectral pattern and concentration, non-invasive methods have become more frequently used. First introduced for monitoring during surgery, they have lately been used to provide hemoglobin spot checks in primary care and blood donor qualification.
Some noninvasive devices use pulse oximetry, while others rely on white light and transmission data capture to measure hemoglobin concentrations in tissue capillaries.
Occlusion spectroscopy is a noninvasive measurement technology featuring a ring-shaped sensor attached to the subject’s finger [5]. The sensor temporarily ceases blood flow, initiating an optical signal that yields a high signal-to-noise ratio. This provides a measurement of hemoglobin concentration.
Sahli´s Method
Sahli’s hemoglobinometer is a manual device that contains a hemoglobin tube, pipette, stirrer, and comparator. Hydrochloric acid converts hemoglobin to acid hematin, diluting until the solution’s color matches that of the comparator block.
The clinician can then ascertain the hemoglobin concentration by reading from the calibration tube. Although this is one of the most common methods for estimating hemoglobin in developing countries and is relatively inexpensive and straightforward, the results are not always precise. For example, there is often inter-observer variability, and it is also highly prone to errors due to manual pipetting [3].
Hematology Analyzer
An automated hematology or hemoglobin analyzer is commonly used to provide high throughput for analyzing various red and white blood cells, hematocrit, and hemoglobin levels from a blood sample. These analyzers offer higher precision values at a fraction of the time compared with manual methods.
The initial cost of an automated analyzer is high, and regular maintenance and the laboratory personnel needed for the device can increase costs. Additionally, stable climate conditions are required, making it an unsuitable option for non-laboratory environments like mobile blood donor test sites and anemia screening projects out in the field.
Blood Gas Analyzer (BGA)
Typically used with arterial blood, BGA measures the combination of blood gas, pH, electrolytes, and metabolite parameters. Some laboratories may use BGAs for hemoglobin testing, but they are more commonly seen in critical care units, delivery wards, and emergency rooms.
The recent development of ready-to-use sensors and solution cassettes with automated calibrations has made BGAs more user-friendly and robust, but maintenance is still required. With the latest innovation of hand-held devices operating single-use cartridges, BGAs have also become available for use in mobile settings.
Anemia is a condition in which the number of healthy red blood cells or the availability of hemoglobin falls below the body’s physiologic needs.
Hemoglobin or hematocrit testing is the primary blood test for diagnosing anemia, which can be caused by poor nutrition or various diseases.
References:
- [1] Shah VB, Shah BS, Puranik GV. Evaluation of non-cyanide methods for hemoglobin estimation. Indian J Pathol Microbiol. 2011;54(4):764-768.
- [2] Vanzetti G. An azide-methemoglobin method for hemoglobin determination in blood. J Lab Clin Med. 1966;67(1):116-126.
- [3] Srivastava T, Negandhi H, Neogi SB, Sharma J, Saxena R. Methods for Hemoglobin Estimation: A Review of “What Works.” Journal of Hematology & Transfusion. 2014.
- [4] van Assendelft OW, Horton BR, Parvin RM. Calibration and control in haemoglobinometry. Clin Lab Haematol. 1990;12 Suppl 1:31-42.
- [5] Pinto M, Barjas-Castro ML, Nascimento S, et al. The new noninvasive occlusion spectroscopy hemoglobin measurement method: a reliable and easy anemia screening test for blood donors. Transfusion. 2013;53(4):766-769.
- [6] EKF Anemia Portal – https://www.ekfdiagnostics.com/anemia-and-hemoglobin-testing.html
