2Dilutions for the Clinical Laboratory Dilution = making weaker solutions from stronger onesExample: Making orange juice from frozen concentrate. You mix one can of frozen orange juice with three (3) cans of water.
3Dilutions for the Clinical Laboratory (cont’d) Dilutions are expressed as the volume of the solution being diluted per the total final volume of the dilutionIn the orange juice example on the previous slide, the dilution would be expressed as 1/4, for one can of O.J. to a TOTAL of four cans of diluted O.J. When saying the dilution, you would say, in the O.J. example: “one in four”.
4Dilutions for the Clinical Laboratory (cont’d) Another example:If you dilute 1 ml of serum with 9 ml of saline, the dilution would be written 1/10 or said “one in ten”, because you express the volume of the solution being diluted (1 ml of serum) per the TOTAL final volume of the dilution (10 ml total).
5Dilutions for the Clinical Laboratory (cont’d) Another example:One (1) part of concentrated acid is diluted with 100 parts of water. The total solution volume is 101 parts (1 part acid parts water). The dilution is written as 1/101 or said “one in one hundred and one”.
6Dilutions for the Clinical Laboratory (cont’d) Notice that dilutions do NOT have units (cans, ml, or parts) but are expressed as one number to another numberExample: 1/10 or “one in ten”
7Dilutions for the Clinical Laboratory (cont’d) Dilutions are always expressed with the original substance diluted as one (1). If more than one part of original substance is initially used, it is necessary to convert the original substance part to one (1) when the dilution is expressed.
8Dilutions for the Clinical Laboratory (cont’d) Example:Two (2) parts of dye are diluted with eight (8) parts of diluent (the term often used for the diluting solution). The total solution volume is 10 parts (2 parts dye + 8 parts diluent). The dilution is initially expressed as 2/10, but the original substance must be expressed as one (1). To get the original volume to one (1), use a ratio and proportion equation, remembering that dilutions are stated in terms of 1 to something:______2 parts dye = ___1.0___10 parts total volume x2 x =x =The dilution is expressed as 1/5.
9Dilutions for the Clinical Laboratory (cont’d) The dilution does not always end up in whole numbers.Example:Two parts (2) parts of whole blood are diluted with five (5) parts of saline. The total solution volume is seven (7) parts (2 parts of whole blood + 5 parts saline). The dilution would be 2/7, or, more correctly, 1/3.5. Again, this is calculated by using the ratio and proportion equation, remembering that dilutions are stated in terms of 1 to something:__2 parts blood_____ = ___1.0___7 parts total volume x2 x =x =The dilution is expressed as 1/3.5
10Dilutions for the Clinical Laboratory (cont’d) Dilution Factor – used to correct for having used a diluted sample in a lab test rather than the undiluted sample. The result (answer) using the diluted sample must be multiplied by the RECIPROCAL of the dilution made.The RECIPROCAL of a 1/5 dilution is 5.
11Dilutions for the Clinical Laboratory (cont’d) Correction for using a diluted sampleExample: A technician performed a laboratory analysis of patient’s serum for a serum glucose (blood sugar) determination. The patient’s serum glucose was too high to read on the glucose instrument. The technician diluted the patient’s serum 1/2 and reran the diluted specimen, obtaining a result of 210 g/dl. To correct for the dilution, it is necessary to multiply the result by the dilution factor (in this case x 2). The final result is 210 g/dl x 2 = 420 g/dl.
12Dilutions for the Clinical Laboratory (cont’d) Sometimes it is necessary to make a dilution of an existing solution to make it weaker.Example: A 100 mg/dl solution of substrate is needed for a laboratory procedure. All that is available is a 500 mg/dl solution of substrate. A dilution of the stronger solution of substrate is needed.
13Dilutions for the Clinical Laboratory (cont’d) To make a weaker solution from a stronger one, use this formula:V1 x C1 = V2 x C2Example: To make 100 ml of the 100 mg/dl solution from the 500 mg/dl solution needed in the previous example:V1 = 100 ml V2 = V2 (unknown)C1 = 100 mg/dl C2 = 500 mg/dl100 ml x 100 mg/dl = V2 x 500 mg/dlV2 = 20 mlDilute 20 ml of 500 mg/dl solution up to 100 ml with water to obtain 100 ml of 100 mg/dl substrate solution
14Serial DilutionsDilutions can be made singly (as shown previously) or in series, in which case the original dilution is diluted further. A general rule for calculating the dilution of solutions obtained by diluting in a series is to MULTIPLY the original dilution by subsequent dilutions.
15Serial Dilutions (cont’d) Example of a serial dilution:
16Serial Dilutions (cont’d) In the serial dilution on the previous slide, 1 ml of stock solution is mixed with 9 ml of diluent, for a 1/10 dilution. Then 1 ml of the 1/10 dilution is mixed with another 9 ml of diluent. The second tube also has a 1/10 dilution, but the concentration of stock in the second tube is 1/10 x 1/10 for a 1/100 dilution.
17Serial Dilutions (cont’d) Continuing with the serial dilution, in the third tube, you mix 1 ml of the 1/100 dilution from the second tube with 9 ml of diluent in the third tube. Again you have a 1/10 dilution in the third tube, but the concentration of stock in the third tube is 1/10 x 1/10 x 1/10 for a 1/1000 dilution.This dilution could be carried out over many subsequent tubes.
18Serial Dilutions (cont’d) Serial dilutions are most often used in serological procedures, where technicians need to make dilutions of patient’s serum to determine the weakest concentration that still exhibits a reaction of some type. The RECIPROCAL of the weakest concentration exhibiting a reaction is called a “titer”.
19Serial Dilutions (cont’d) Example of determining a titer:A technician makes a serial dilution using patient serum:Tube #1 = 1/10Tube #2 = 1/100Tube #3 = 1/1000Tube #4 = 1/10,000Tube #5 = 1/100,000Reactions occur in tubes 1 through 3, but NOT in tubes 4 or 5. The titer = 1000.