Measured Low Density Lipoproteins (LDL)
Cholesterol is part of the family of lipids, more commonly known as "fat." It is a fatty substance that circulates in the blood and is essential for the good functioning of the body.
It participates to the construction of the cell membrane, ensures the smooth flow of cells and their permeability by certain substances. It is also involved in the production of bile salts whose role in digestion is essential, as well in the synthesis of many hormones such as sex hormones including progesterone or testosterone. Cholesterol is transported in the blood by proteins, lipoproteins.
Lipoproteins are usually classified according to their size and density, with chylomicrons, chylomicron remnants, and very low density lipoprotein (VLDL) being relatively large and light, whereas LDL and HDL are consecutively smaller and heavier. The size of LDL particles varies such that particles with more triglycerides and fewer cholesteryl esters result in smaller denser LDL.
There are two main types of lipoproteins.
LDL transports cholesterol from the liver to the organs that need it. This is called “bad cholesterol” since it has been linked to an increased risk of developing cardiac diseases.
HDL does the opposite direction: they recuperate excess cholesterol in the tissues and carry it back to the liver, where it is "recycled" into bile acids. It is called “good cholesterol” since it removes excess of cholesterol from the vascular system preventing atherosclerosis.
Many risks of cardiovascular diseases are associated with LDL; this macromolecule is considered undesirable because it deposits excess cholesterol in the walls of blood vessels and contributes to hardening of the arteries, and the development of heart disease.
LDL deposited in the arteries accelerates the
formation of plaques. So it induces obstruction of the blood circulation and consequently the inhibition of the oxygen supply and nutrients to the cells, tissues and organs.
Atherosclerosis, indeed, refers to the presence of plaque on the inner walls of the arteries which obstructs partially or completely the flow of blood. This is a very slow process and it may take many years before an attack of cardiac or other symptoms occur. Atherosclerosis mainly affects large and medium-sized arteries (eg, coronary arteries, the arteries of the brain and the arteries of the limbs).
This phenomenon is observed in patients with metabolic syndrome and characterized by elevated plasma triglycerides (150mg/dL or 1.7mmol/L), a reduced cholesterol content of high density lipoprotein (HDL) cholesterol (<40mg/dL or 1.0mmol/L for men and <50mg/dL or 1.3mmol/L for women), an increase of a subclass of small, dense LDL particles (>160mg/dL), depleted of cholesterol. 
Reducing levels of small, dense LDL particles may counteract the risk of cardiovascular diseases.
The test measures the amount of LDL cholesterol in whole blood collected from subjects with or without food and is respectively evaluated by two different methods.
It is obtained applying the Friedewald equation:
LDL-C = TC-HDL-TG /5.0 (mg/dL)
This formula is indeed based on the concentrations of total cholesterol (TC), High Density Lipoprotein of (HDL) and the concentration of triglycerides (TG). 
Consequently the test is dependent of some restrictions, since the blood sample has to be drawn in fasting state to limit interference of triglycerides.
The reference method to directly measure of LDL is based on the physical properties of these macromolecules. It requires ultracentrifugation of serum (5 mL) at its own density (1.006 g/mL) for 16 h at 120 000g, with the removal of triglyceride-rich lipoproteins in the supernatant fraction. However this is not feasible in clinical practice and another procedure has been developed for the laboratory routine.
The current method to have a direct LDL is to measure serum or plasma cholesterol concentrations via enzymatic analyses. This method provides results that are very comparable with those obtained with the reference method and can be performed by automated clinical chemistry systems. 
Moreover, this method to directly quantify the concentration of low density lipoprotein is independent from the HDL and TG values, so that patients do not need to be fasting.. Therefore LDL can be directly measured in subjects at any time of day and in all circumstances (emergency or routine) without that nutritious factors, due to recent meal, counterfeit the result.
The amount of cholesterol in the blood is measured in order to predict the risk of development of cardiovascular disease. LDL is considered as one of the most relevant marker for the prediction of the cardiac and metabolic disease. Indeed therapeutic decisions are often based on its measurement.
In order to identify patients at risk of cardiovascular disease by the detection of large amounts of cholesterol, LDL-C measurement seems appropriate because it allows having a total lipid profile. Thus the clinician with an overall idea of the lipid concentrations can more easily decide on appropriate therapy. However the Friedewald formula may not reflect the real values of LDL especially when triglyceride values are high (≥150 mg/dL) or when the HDL values are also very elevated. Since during the day triglyceride levels fluctuate on average, 0.5 mmol/L in women and 1.0 mmol/L in men, the LDL-C is affected by this variation, with the risk of underestimation when measured in the non-fasting state.
This underestimation can increase when untreated LDL concentrations are very low or when LDL is aggressively lowered by potent statins and other lipid-lowering interventions. Similar under estimation of true concentrations of LDL can also occur when HDL concentrations are very high, an issue that has been reported in some studies of inhibitors of cholesteryl ester transfer protein (CETP).
Based on these facts, high-risk patients should have additional evaluation, especially if triglyceride levels are ≥150 mg/dl. [4, 5]
One of the keys to overcome these problems is to use the values of the direct measurement of LDL in order to have a reliable result which reflects the accurate value of LDL in the blood.
Compared with LDL-C, LDL-D measures could also be better for assessment of very low concentrations of LDL after aggressive reduction with therapy. Moreover, in diabetic patients, overweight patients and non-fasting who consumed large quantities of fatty foods, direct measuring LDL is most appropriate. 
< >Tietz Fundamentals of Clinical Chemistry and Molecular Diagnostics, 7e (Fundamentals of Clinical Chemistry (Tietz... by Carl A. Burtis PhD and David E. Bruns MD (May 8, 2014)Friedewald WT, Levy RI, Fredrickson DS. Estimation of the concentration of low- density lipoprotein cholesterol in plasma without use of preparative ultracentrifuge. Clin Chem 1972;18:499–502.Otokozawa S, Ai M, Asztalos BF,Schaefer EJ et al. Direct assessment of plasma low density lipoprotein and high density lipoprotein cholesterol levels and coronary heart disease: results from the Framingham Offspring Study.Atherosclerosis. 2010 Nov;213(1):251-5.
< >Davidson M, Liu SX, Barter P, Brinton EA, Cannon CP, Gotto AM Jr, Leary ET, Shah S, Stepanavage M, Mitchel Y, Dansky HM. Measurement of LDL-C after treatment with the CETP inhibitor anacetrapib. J Lipid Res. 2013 Feb;54(2):467-72.Martin SS, Blaha MJ, Elshazly MB, Brinton EA, Toth PP, McEvoy JW, Joshi PH. Friedewald-estimated versus directly measured low-density lipoprotein cholesterol and treatment implications. J Am Coll Cardiol. 2013 Aug 20;62(8):732- 9.Lund SS, Petersen M, Frandsen M, Smidt UM, Parving HH, Vaag AA, Jensen T. Agreement between fasting and postprandial LDL cholesterol measured with 3 methods in patients with type 2 diabetes mellitus. Clin Chem. 2011 Feb;57(2):298-308.
In this ever advancing world of cell counters where does a small lab with a 3-part differential cell counter stand, that was my question to myself when I started with Micros 60. I thought okay this the best we got and made some observations over 1-year period. As we all must be aware plasmacytoid lymphocytes are quite characteristic for dengue patients(citation), so I tried sometimes to go retrospective in case of plasmacytoid lymphocytes in confirmed dengue patients and tried looking at flags of Micros, and I found consistent M2 G1 G2 flag in dengue cases with thrombocytopenia, after sometime I become confident of this flag and any patient with classical high grade fever , calf tenderness and raised sgot sgpt enzyme levels having this flag on CBC would make me smile like a magician on being sure that it is dengue and I will get plasmacytoid lymphocytes in smear. But these observations are not true for every case, as I also observed similar flags in leukaemia and cases of acute hepatitis as other people (citation), so I suggest that in country like ours which sees epidemics of dengue year after year incorporating this M2 G1 G2 flag in screening algorithm for dengue will help us in detecting more cases of dengue during outbreaks.
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