Mod3 Hema Lec- anticoagulation ADDITIVES.docx.pdf

HEMATOLOGY 1 LECTURE | 3RD YEAR | FIRST SEMESTER BATCH SORBETES ‘25 MODULE 3: ANTICOAGULANTS & ADDITIVES ANTICOAGULANTS...

HEMATOLOGY 1 LECTURE | 3RD YEAR | FIRST SEMESTER BATCH SORBETES ‘25 MODULE 3: ANTICOAGULANTS & ADDITIVES ANTICOAGULANTS 1. EDTA (ETHYLENEDIAMINETETRAACETIC ACID) Prevent blood from clotting by mechanisms that ⟾ This is the anticoagulant of choice for MOST disrupt the Coagulation Cascade hematologic test ⟾ Basta hematology, the majority of hematologic test will ⟾ The medical jargon of clotting is coagulation, meaning be Lavender top or EDTA.So if you don’t know or not it's the same as clotting. Clotting is just a simplified sure na nasa Hema siya, EDTA agad isagot. Under ng form but the technical term for that is coagulation. hema, #1 na hahanapin will be EDTA. Kahit hindi niyo ⟾ From the term “ANTI”, kalaban niya ang coagulation. It na talaga alam. Of course Blue top is under hema, prevents the coagulation cascade. special coagulation test. ⟾ Coagulation cascade is actually cascaded by coagulation factors. a. Used in the concentration of 1.5mg/mL of blood ○ Factor 1,2,3,4,5,7,8,9,10,11,12,pkh b. Mechanism – Chelates Calcium; This process forms ⟾ If there is lacking or you prevent one of the coagulation an insoluble calcium salt that prevents blood factors. The coagulation cascade will be inhibited or coagulation. will not proceed. So in simpler terms ganun yung i. When you say chelates, synonymous siya sa to mechanism ng ANTICOAGULANT. bind. So it simply binds with calcium ○ They disrupt the coagulation cascade by ii. Why calcium? Anong meron sa kanya? inhibiting the coagulation factors. That initiate Because calcium is a coagulation factor IV, or w/c are the component of the coagulation makikita sa text books/literature, hindi Factor IV cascade kundi Calcium pero same as factor IV ⟾ ADDITIVES iii. Tinanong seaboard exam Last yr, which is a ○ When you say additives are they the same as mineral coagulation factor? The mineral anticoagulants? or are anticoagulants additives? coagulation factor is FACTOR IV or CALCIUM. Or are all anticoagulants additives? anong meron sa kanya? Because calcium is Yes involved in the 3 pathways of coagulation ○ Are additives always anticoagulants? cascade. You have extrinsic, intrinsic, and No, kasi example is may sodium fluoride na common pathways. andun lahat si calcium. gray top tube which preserves glucose. It That's why if you inhibit the calcium – (lag si sir) doesn't anticoagulant blood lahat ng anticoagulants are additives BUT c. Anticoagulant of choice for hematology counts not all additives are anticoagulants. because it maintains good morphology By the name itself additives, ena-add mo i. choice for hematology, pinaka importante na lang siya but it doesn't necessarily mean malaman. Lahat or almost, majority of that this are anticoagulants. hematologic test is EDTA because it maintains The example by Ms. Rosario :), sodium good morphology fluoride which is NOT an anticoagulant, it is an additive that preserves glucose. There is d. Forms: a separate additives that prevents i. Disodium Salt (Versene) coagulation cascade na nasa gray tube ii. Dipotassium Salt (Sequestrene) ○ The gel in the tube are additives (anticoagulant iii. Tripotassium Salt which is usually in liquid ba siya? Hindi) form ○ Clot activator is not also an anticoagulant but considered as additives ***The amount of EDTA per milliliter of blood is essentially ○ ALL ANTICOAGULANTS ARE ADDITIVES. Kasi the same for all three forms of EDTA. ena-add mo sila, as such under siya ng umbrella Anong amount? 1.5mg/mL of blood, regardless of the ng additive. form kung Versene, Sequestrene, or Tripotassium Salt ○ HUWAG sasabihin na ALL ADDITIVES ARE ALWAYS ANTICOAGULANTS because you are also adding solutions/substances that are not anticoagulants 1 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 SIDENOTES: PLATELET SATELLITOSIS Excess EDTA causes shrinkage of red cells A rare condition that occurs when an IgG antibody resulting to decreased Hct and ESR forms in the presence of EDTA, the anticoagulant ○ Pag sumobra sa EDTA, dba 1.5mg/mL of blood, if you that is used for the collection of hematology blood have excess EDTA for example short draw (may na specimens. The IgG antibody is directed against draw pero gumalaw and na miss ulit), in this case there the glycoprotein lIb/Illa complex on the platelet will be excess EDTA. Anong mangyayari? shrinkage of membrane. As the antibody coats the platelets, the red cells resulting to decreased Hct and ESR platelets rosette around segmented neutrophils, Hct is also known as Packed Cell Volume (PCV) – bands, and sometimes around monocytes. packed na sisidsid. So kung maliliit yung RBC, From the term itself again, Platelet; satellite. your Hct is low. Yung tube na pinakita ni doc, APara siyang asteroid na nag satellite sa Jupiter. which is diveided into liquid and solid, the solid Sino si jupiter? Si neutrophil, si bands, and component excluding the buffy coat is the Hct. And monocytes. They satellite around neutrophiles, the bulk of that solid phase ay RBC. So if maliliit bands, or juvinile neutrophils, and or monocytes RBC mo, expect the lvl ng solid phase mo is low sometimes. Bakit? Kasi if EDTA is used, there is erythrocyte sedimentation rate (ESR) – from the a rare condition that occurs when an IgG term itself, erythrocyte (RBC), sedimentation (yung antibody forms in the presence of EDTA. pagbaba nila), rate (gaano kabilis or kabagal yung Anong pangalang ng antibody na iyon? anti pag settle sa baba ng tube yung mga erythrocytes Glycoprotein (GP) IIb-IIIa Isipin/imagine niyo, if maliliit yung mga RBCs magagaan sila, if magagaan yung mga GP IIb-IIIa will be seen in the surrounding platelet. The RBCs, hindi sila mabilis bumababa or nag cell in our body have receptors, for signal, parang antena. ssettle down. At dahil diyan decrease ang Isang antena makikita mo sa membrane ng platelet ang ESR. Kasi masyodo silan magagaan kasi GP IIb-IIIa. It is an athena or receptor for fibrinogen. shrink or maybe too small Itong GP IIb-IIIa, may na form daw na antibodies against ESR is used to indicate inflammation. If it if EDTA ang ginamit and dahil dito yung mga antibody mataas ang ESR, you can say that there is nayun will coat the platelet. And the platelet will rosette an inflammation around segmented neutrophils, band, and sometimes The suffix -itis attached after an organ, monocytes. simply means inflammation Actually asked in the board exam TWICE. San sila nag So if doc asked, appendicitis is a. infection; b. rrousette or ano ang cause ng platelet satellitosis? inflammation. kahit bacteria pa ang cause ng ANTIBODY appendicitis, if the suffix -itis is attached after the organ, it The next question after that boarded exam na, naulit but means inflammation, regardless of its etiologic cause diff. question. What is the antibody formed during platelet kung bacteria or viral yan satellitosis? anti Glycoprotein (GP) IIb-IIIa - Say hepatitis (VIRAL), the suffix -itis attached after an organ simply means inflammation Antibody-coated platelets that are huddled around white blood cells (WBCs) will not be counted as platelets by In ESR hindi masyadong ginagamit ngayon kasi meron automated equipment and the platelet count will be na tayong more specific indicator for inflammation. That’s falsely decreased. If a peripheral blood smear is why as a medtech, you would know yung doctor na yun is reviewed, platelets will be observed attached to WBCs hindi Medtech if nag order siya ng ESR (diff. premed) kasi (labce.com). hindi deep yung alam niya sa ESR. Advantage nating What is the effect on platelet count? so yung platelet Medtech if mag MD is alam niyo ang test/updates. ESR count if gagamit ka ng automated instrument, ma account ay hindi specific ba siya as platelet? Hindi na, kaya mag kakaroon ng falsely decrease of platelet. Nasaan ba kasi yung mga platelet? nag ssurround na sila or parang nakadikit na sila sa mga WBCs, kaya hindi sila na ccount individually as platelets. AS such there is false decrease ng platelet count. And if you will see it at peripheral blood smear as a medtech, makikita mo na may maliliit na parang tipak na bato that is naka surround sa isang malaking bato PLATELET SATELLITOSIS Paano makikita sa question? Low platelet then medtech sumilip under microscope then nakita niya na may mga nakadikit na particles/fragments na nag ssatellite or surrounding a bigger cell. Ito ay PLATELET SATELLITOSIS cause by IgG antibodies directed against GP IIb-IIIa receptors found in the membrane of platelets 2 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 Kung ang tanong naman is anong ang gagawin mo if 2. CITRATE mababa ang platelet count pag tingin mo sa microscope Pareho lang sila ni EDTA. Majority of the anticoag. ang nung gumawa ka ng PBS nung eni-stain mo nakita mo kanilang mechanism is that they will bind with calcium in may platelet satellitosis, discard ba yung spx or recollect a soluble complex. Insoluble kanina pero dito according ka? Mag RECOLLECT ka. perform another venipuncture kay Steininger, soluble complex Gagamit ka ba ng the same anticoagulant? YES, Binds Calcium in soluble complex gagamit ng ibang anticoag. alam mo na nga yung Anticoagulant for PNH (Paroxysmal Nocturnal cause uulitin mo pa, DO not repeat the same mistake Hemoglobinuria) testing (will be discuss in IS pero pag twice sa lab. Instead of EDTA, use sodium citrate ttest mo siya, under ng Hema. Although magiging topic (blue/light blue top). However, REMEMBER, iba yung sa IS) volume ng citrate sa volume used in EDTA. Because of ○ To give a birds eye view, in PNH there is deficiency the diff. of the dilution of blood in relation to the anticoag. of regulatory protein for complement you have to correct the platelet count. How to correct it? ○ Deficiency of decay accelerating factor (DAF) - By multiplying the platelet count by 1.1. Why? also known as CD55 (class designation or cluster of kasi nag karoon ng diff of dilution, kaya iba ang differentiation 55) dilution ng blood. To correct it, isalng mo dun sa DAF/CD55 regulates the complement para machine, if nag appear yung platelet count, hindi sobra. Kasi if na activate ang complement huwag yun e-report mo, multiply muna sa proteins ang end result is lysis. If you did not obtained platelet count by 1.1 na galing sa control or regulate the complement, it will be machine and then yun yung e-report mo as result continuous of lysis of patient So kung low or walang DAF, walang mag rregulate ng lysis The phenomenon of “platelet satellitosis” may occur when using EDTA anticoagulants. platelets adhere Ratio of blood to anticoagulant is 9:1 around neutrophils, forming a ring or satellite effect. ○ 9 parts of your blood vs 1 part of anticoagulant Using sodium citrate as an anticoagulant should ○ What is the meaning of this? How many mL or what correct this problem. Because of the dilution in the is the volume of blood to be drawn in blue top citrate tubes, it is necessary to multiply the obtained containing 0.3 mL of citrate? what if tinanong ni sir. 9 platelet count by 1.1. is to 1, hindi 9mL by 1 mL, 9 parts of your blood vs 1 part of anticoagulant ○ For example, kadalasan laman ng light blue top is 0.3 mL. Ilan mL or volume of blood should you get or mix to tube? By ratio and proportion, 9 x 0.3 = 2.7 so how many mL ang ilalagay sa light blue top is 2.7 mL of blood. basta remember 9:1 kung blood to anticoag ratio Example a view under a microscope, yung naka circle na malaki is RBC; maliit, a central pallor occupying ⅓ of the diameter of RBC. Concentrated yung Hb sa periphery. Mag focus dun sa purple color, which is a WBC particularly a neutrophil. Yung maliliit surrounding it is What if nilito tayo ni sir, anticoag to blood platelets (rosette), at least may makikita kang tatlo. That ratio; 1:9 will already form or considered as rosette. Anong affect pwede mag vary depende kung ano ibigay na niya sa platelet count? Falsely decreased value. Basta REMEMBER: 9 is to 1 blood to RESOLUTION? perform another venipuncture using anticoagulant ratio or 1 is to 9 anticoag to blood ratio another anticoag. BUT multiply your platelet count by a Hindi yung lagay lang ng lagay since it is factor of 1.1 due to differences of dillution crucial, sa pt appt or coagulation test sa hema, crucial ang anticoag. to blood ratio or vice versa 2 Forms: ○ Disodium Citrate (black top) ESR ESR to be discussed midterms or finals Hindi porket citrate ang gamit mo sa black top 9 is to 1 din ang ratio. HINDI, sa light blue top lang applicable ○ Trisodium Citrate (light blue top) 3 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 Coagulation and platelet function studies Conflict siya kay Rodak, kasi sabi ni Steininger 50%, (which is also under hematology section pero dahil competitive tayo nagbasa tayo ng Rodaks because that is hema2. NOT ALL the time because this is our mother reference. Steininger 50 % EDTA ang isasagot. We also have light blue and Rodak 55%. top) Preserves the labile factors V and VIII ○ The 9:1 blood-to-anticoagulant ratio is effective Itong V and VII kasi, mentioned kanina provided the patient's hematocrit is 55% (or.55 L per yung diff. coag factors. These 2 if Liter) Less. In polycythemia, the decrease in plasma pinatagal yung test, ex. update kay jowa, galit volume relative to whole blood unacceptably raises na siya, kasi toxic na, kaya lang si jowa na ito the anticoagulant-to-plasma ratio, which causes makitid ang utak (malas mo), alam niyang toxic dun falsely prolonged results on clot-based coagulation sa lab tapos dadamay pa siya kaya na delay ka umabot ng 3hrs, the factors V and VIII will tests.(rodak) deteriorate will result to FALSE In polycythemia, there's decrease plasma DECREASE or say deficiency na ng V Nasaan ang coagulation factors decrease? and VIII when in reality wala naman NASA PLASMA, so parang decrease na din siyang factor V and VIII deficiency. ang coagulation factors mo.Magclot ba siya on ROLE OF CITRATE IS TO PRESERVE. Kasi madali itong time? HINDI, kaya prolonged ang clotting time mawala or deteriorate if test is not done ASAP. And this is the UNIQUE advantage of citrate kung bakit siya Na pwedeng masabi mo “Uy may coagulation ginagamit sa coagulation and platelet function studies factor deficient tong patient na to, baka may hemophilia siya” HINDI, MALAY MO MAY Most satisfactory for platelet aggregation POLYCYTHEMIA PALA YONG PATIENT. Not studies necessarily hemophilia siya, okay? okay. SIDENOTES: So ito yong resolution, the formula is: ★ Black-top tubes also contain buffered sodium citrate C = (1.85×10^-3 )(100 -H)V; and are generally used for Westergren sedimentation ○ where C is the volume of sodium citrate in milliliters, V rates, as are lavender-top tubes. They differ from light is volume of whole blood-sodium citrate solution in blue-top tubes in that the ratio of blood to milliliters, and His the hematocrit in percent (Rodak) anticoagulant is 4:1 in the blacktop tubes and 9:1 in If a patient is polycythemic, let’s say yong the light blue-top tubes. hematocrit niya 65% or. 65L per liter then ★ If the Hematocrit value exceeds 0.50L/L, as in prolong ang kanyang PT and APTT your Polycythemia, or if there is incomplete filling of the resolution will be this one: C = (1.85×10^-3 )(100 sample tube, the amount of unbound citrate in the -65)V. The volume diba 2.7 plus 0.3, so that’s citrate:plasma mixture causes a prolongation of 3mL, that’s the total volume of the blood and the clotting times, particularly in PT and APTT tests for anticoagulant, now: clotting times. (Steininger) C = (1.85×10^-3 )(100 -65)V. What if mataas ang Hct ng patient? Again paano muna si = (1.85×10^-3 )(35) x 3 Hct, yung PCV in relation to liquid phase (plasma), = 0.19mL then round to 0.2mL (gagamitin mong according Steininger if nag exceed daw ng 0.50L/L or volume ng anticoagulant para hinid ka sumobra 50% yung kanyang Hct as in polycythemia, malapot yung sa citrate or anticoagulant. Instead na 0.3 blood niya or RBC count is mataas, kaya tumataas din bawasan mo ng 0.1 *laughs*, 0.2mL ang gamitin PCV na kapag nag exceed daw ng 50% or 0.50L/L there mo, okay? Para prevent yong excess unbound would be incomplete filling of the sample tube citrate. Bakit falsely prolonged? Hct increased, means plasma is ○ Although 3.8% sodium citrate solution is still used, displaced. Ano ang sinabi ni sir last time? na saan yung 3.2% buffered sodium citrate has been adopted as coagulation factors? Nasa PLASMA, kung na displace the standard anticoagulant for coagulation studies by yung plasma tumataas yung Hct, kumukonti yung plasma. the National Committee for Clinical Laboratory At dahil kumokonti yung plasma kasi ino-occupy iyon ng Standards (NCCLS) and College of American packed RBCs, what will happen? Parang kumonti na rin Pathologists as well ang coag factors, and with these agsursurok or Although 3.8% sodium citrate solution is still sumosobra ang citrate. there is unbound excess in the used, 3.2% buffered sodium citrate both will be citrate at dahil dyan sobra sobra or parang there is a correct but if you will be up to choose only one leakage of plasma the prolonged yong coagulation or 3.2 percent. Basahin mo para intense. yong clotting time or both PT and APTT. 4 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 3. OXALATE (DOUBLE OXALATE) Thrombin is factor 2A or activated prothrombin because thrombin is the inactivated form. So, Used in the concentration of 2mg/mL of blood pag ininhibit mo to, anong manyayari? Walang Binds calcium to form insoluble calcium oxalate coagulation cascade. Anticoagulant for PNH testing (deficiency of May be the anticoagulant of choice when blood is what? decay accelerating factor (DAF) or CD55) collected for the platelet retention test (glass bead anong tawag sa test na yon? Serum hemolysis retention test) Test. Used to coat capillary blood collection tubes. Forms: Yong red band, may anticoagulant may ○ a.Ammonium Oxalate anticoagulant yon pero ang blue band walang Swells the red cells anticoagulant yon. Diba sa mga stoppers natin pag ○ Potassium Oxalate red top walang anticoagulant? pero pag sa capillet Shrinks the red cells natin meron. Yong anticoagulant sa red band is the HEPARIN. EDTA ilan? 1.5 milligram per ml. Excess EDTA would An inappropriate anticoagulant for many hematology cause shrinkage in RBCs, right effects sa hematocrit tests, including Wright-stained blood smears as it gives a bababa, effects sa ESR mababa, right? Okay balik tayo blue background with Wright stain. (TANDAAN NIYO sa oxalate, ganun pa rin guy, binds calcium to form TO) Sa ibang pages ni Rodak sabi niya gray or dark insoluble calcium oxalate.Although sa ibang reference, gray. Kay Henry naman, bluish ang background niya citrate diba kanina sabi ko soluble pero sa ibang kaya hindi mo pwedeng gamitin ang blood na may reference insoluble. Suffice to know nalang that all three heparin as anticoagulant kapag gumagawa ka ng blood EDTA, Citrate, and Oxalate or bind calcium to inhibit smear or peripheral blood smear. coagulation cascade. Ano ba kasi ang nagprepreserve ng mga morphology ng mga cells? Sinabi kanina ni Sir, Saang stopper si Oxalate? Sa gray top. The EDTA. Kaya pag nagbloodsmear gumamit ng purple anticoagulant present in gray top is oxalate. Although top, wag iba kasi bibigyan ka ng ng blue background present din doon si EDTA pero depende sa brand or with wright stain. depende sa manufacturer, but the anticoagulant Heparin has an advantage over EDTA as an present together with the sodium fluoride is your anticoagulant, in that it does not affect levels of ions such oxalate.Although may mga ibang manufacturer as calcium. However, heparin can interfere with some nagdagdag sila ng EDTA instead of oxalate. immunoassays. Heparin should not be used for 4. HEPARIN coagulation or hematology testing. Heparinized plasma is preferred for potassium measurements to avoid an used in the concentration of 15-20U/mL elevation due to the release of potassium from platelets Uy, para saan si Heparin? Anong sinabi ni Sir last as the blood clots. time sa arterial puncture? For arterial blood gas Heparin has an advantage over EDTA as an analysis. Pag narinig niyo sa hospital ABG ABG anticoagulant, in that it does not affect levels of ABG, that’s arterial blood gas analysis. ions such as calcium. Bakit? Kasi ano ba ang action ng heparin? Heparin descalate bind with Ano naman ang concentration niya paggamit mo ang calcium, your calcium will be unaffected, compare to ABG? Anong sinabi ni Sir last time? That's 1000 units your EDTA it collates or bind with calcium as such per mL. Anong sabi ni Bishop, most likely or most of the your ionized calcium will be falsely decrease. time or generally excess heparin is the most common cause or pre-analytical error, naalala niyo from our Heparinized plasma is preferred for potassium specimen collection lecture? Right. measurements to avoid an elevation due to the Natural anticoagulant (in vivo anticoagulant) and in vitro release of potassium from platelets as the blood anticoagulant. clots. Bakit ano ba ang ginagamit natin for potassium Acid mucopolysaccharide that inhibits coagulation by measurement? Part siya ng chemistry, electrolytes. inactivation of thrombin and Factor Xa (Turgeon, Rodak) Kapag chemistry section ang request ng doctor, red Is it proper to call it heparin? NO, as an acid top. But for potassium measurement, heparinized mucopolysaccharide this is known as HEPARAN plasma is preferred to avoid an elevation due to the SULFATE, okay? Kasi parang ito yong original na release of potassium from platelets as the blood form ni heparin. clots. Diba kasi sa red top wala syang anticoagulant magclot talaga, pag nagclot kasi si platelets By the way, your thrombin is also known as irereleased niya si potassium, so saan mapupunta FACTOR 2a. Pag may nakita kayong subscript na A su potassium? Sa serum, ano bang sample? for or katabi niya A, ibig sabihin activated enzyme. So potassium. Since napunta siya sa potassium kung 10 lang siya ibig sabihin inactive siya pero elevated or falsely elevated ang potassium kapag 10A ibig sabihin activated na siya, ongoing na ang coagulation cascade. 5 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 Lithium heparin may be used for most chemistry tests NOTE: Heparin increases the RATE of factor except for lithium and folate levels; for lithium, a serum inactivation, not the magnitude. (Hindi niya paLalakasin specimen can be used instead. yung ability of antithrombin 3 through inhibit coagulation Except for lithium, bakit? Syempre may laman cascade. It will only accelerate the reaction.) Rate is siyang lithium tas mag magmemeasure ka ng accelerated from 2,000 to 10, 000x that when AT-Ill alone lithium, ano yon niloloko mo sarili mo? neutralizes Thrombin. Hindi niya ini-increase yung magnitude or the strength or Or lithium, a serum specimen can be used instead. the intensity or the power. It only accelerate the overall Sabi ko nga kung wala kayong ideya pero alam reaction niyong chemistry test yon, red top. 5. MISCELLANEOUS(antiglycolytic agents and separator gel/thixotropic gel) -Sodium heparin cannot be used for assays measuring sodium levels, but it is recommended for trace elements, a. ANTIGLYCOLYTIC AGENTS (From the term itself. leads, and toxicology. Sodium heparin is the injectable Anti; Glycolysis. Anti molism? of glucose form used for anticoagulant theraphy. i. Inhibits glycolysis; for glucose determination Naglolokohan na naman tayo, malamang meron since it preserves glucose. HOW? By siyang sodium, tapos imemeasure mo sodium? Ano inhibiting glycolysis. CONTAINS: kaya ang effect niya, increase or decrease? Edi 1. Fluoride (NaF) increase kasi parang nagdagdag ka lang ng sodium. 2. lodoacetate ii. Are these two anticoagulants? NO, they are Sodium heparin is the injectable form used for antiglycolytic agents anticoagulant therapy. For example, yong mga iii. IF doc asked, what are the anticoagulant stroke patient diba meron silang coagulation, or found in your gray top nagkakaroon ng infarct kaya need natin iprevent yan 1. A. fluoride by simply isa sa management niya ay anticoagulant 2. B. lodoacetate in the form of heparin sodium na ininject. 3. C. oxalate As an antiglycolytic agents, paano mechanism ni fluoride? SIDENOTES: According to HENRY ★ The heparin molecule is a highly negatively charged SIDENOTES: conjugated polysaccharide. By itself, it has little or no As an antiglycolytic agents, paano mechanism ni fluoride? anticoagulant properties (it is not actually an According to HENRY anticoagulant), but when it combines with antithrombin ★ Fluoride, used for glucose determinations, prevents Ill, the effectiveness of antithrombin Ill for removing glycolysis by forming an ionic complex with Mg++, thrombin increases by a hundredfold to a thousandfold, thereby inhibiting the (magnesium) Mg++- dependent and thus it acts as an anticoagulant. (Steininger) enzyme, enolase. (Henry) But by itself it cannot inhibit or it is not an ○ Your enolase is an enzyme needed in glycolysis. It is effective anticoagulant alone, need niya already in the terminal process or pathway needed in pumartner kay antithrombin 3. ANTI-thrombin 3, glycolisis. common sense na to. ○ And needed ang magnesium, since when you inhibit magnesium its like your also inhibiting enolase Si anti-thrombin 3, hindi lang naman si thrombin ○ Para hindi makalimutan, si enolase, siya ang enzyme ang inihiinhibit niya, a lot of coagulation. na enolase (“inalis”) ni fluoride sa glycolysis According to horizon, it inhibits all coagulation ★ Sodium fluoride prevents glycolysis for 3 days but is factors except factor 7. From other references, it neither adequate nor effective in preserving glucose can inhibit factor 9, 10, and 11. concentration in cases of bacterial septicemia. (Henry) ○ Hindi siya effective if your patient have septicemia or For example, the picture. A schematic of how heparin overwhelming bacterial infection. WHY? Kasi si accelerate the function of antithrombin 3 as anticoagulant/ bacteria, san siya nag cconsume ng energy? GLUCOSE, kaya kahit ma prevent yung glycolysis, ARGININE SITE kung meron naman gumagamit ng glucose na LYSINE SITE bacteria. Walang kwenta. Your antithrombin 3 is actually an enzyme, in general, ang ○ So your sodium fluoride is not applicable to all cases tawag sa kanila, ang mga coagulation factors or proteins of patients. NOT EFFECTIVE in cases of bacterial natin na related sa coagulation is SERINE PROTEASE, septicemia when we say serine protease, sila ay pwedeng ma ○ Effective siya in inhibitng glycolysis, however bacteria activate na enzyme. And when it is activated by an competes with them or they take advantage of the activator/signal/messenger, and binds with arginine site, presence of glucose na priniserve ni sodium fluoride, because of the binding of heparin sa kanyang lysine site, mga bacteria ang gumagamit kaya wala kang na Your antithrombin 3 will be activated mmeasure and if meron man, konti lang 6 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 b. SEPARATOR GEL/THIXOTROPIC GEL i. Inert material that undergoes temporary change in viscosity during the centrifugation SIDENOTES: process resulting to separation of serum or ★ Therapeutic drug monitoring specimens should NOT plasma from the cellular constituents be collected in tubes that contain gel separators ii. Present in Gold, Orange, Red, and Gray Top because some gels absorb certain drugs, causing a Naka kita na raw tayo nito, nakikita ito sa gold top and falsely lowered result. ( In our CC, we will be having orange top and pwede rin may makita sa red top and toxicology, and we have a topic, therapeutic drug pwede rin gray top. Meron silang separator or thixotropic monitoring. Basically, you will monitor the drug if it is gel, from the term itself separator, separates the liquid effective or if madaling na metabolize or sobra sobra, component against the solid components kapag mmonitor mo siya. Of course, DON'T USE GEL cinentrifuge mo siya ng isang beses it will separate na. SEPARATOR because it will falsely lower the result Which is a unique advantage because if transfer mo yung since gels absorb certain drugs. Mas maganda yung patient, lets say (???) at least kapag nakukulog kulog, na walang anticoagulant na wala ring gel separator separate na sila. Unlike in red top or top w/o separator ★ Significant DECREASES in phenytoin, gel, the serum will be mixed, have the tendency of phenobarbital, lidocaine, quinidine, and mixture. Compared sa may separator gel, at least fix na carbamazepine have been reported when stored in yung separation nila. Vacutainer SST tubes. (Henry) ★ Advantages of serum separator tubes include Para hindi makalimutan ano yung mga top that have ○ (1) ease of use, separator/thixotropic gel: oohh you’re GORG (gorgeous) ○ (2) shorter processing time through clot activation, Pwede rin daw G OR G (since they also have activator, thrombin) G - Gold ○ (3) higher serum yield, (Kasi nag separate na talaga O - Orange siya) R - Red ○ (4) minimal liberation of potentially hazardous G - Gray aerosols, (at least may barrier ka, para hindi madagdagan yung aerosols) *** Red/gray- and gold-top tubes contain both clot ○ (5) only one centrifugation step, (kasi kapag na activator and a separation gel. centrifuge mo na,they are separated, they won't mix already Red/gray- and gold-top tubes and even you orange top, aside from the separator gel/thixotropic gel, pwede mo ○ (6) use of single tube (same one as patient siyang haluan ng clot activator, from the term itself specimen), and activate the clot, para mapabilis. Kasi in general, 60mins ○ (7) ease of a single label. ang aabutin, kailangan mo pang antayin yang 60mins or ★ A unique advantage is that centrifuged specimens 30mins para mag separate yung serum dun sa clot. Of can be transported without disturbing the course you can just accelerate the coagulation by simply separation. (Henry) adding a CLOT ACTIVATOR. For example thrombin. ○ Because na separate na sila, fix na yung separation. COmpared sa wala kang separator gel, maalog alog. Tendency may mahalo sa serum. ★ Some silica gel serum separation tubes may give rise to minute particles (piece, fragment, or amount) that can cause flow problems during analysis. Filtering the serum solves the problem. (Henry) ○ So if meron daw silica sa gel mo, it may give rise to minute particles na pwedeng ma suck ng machine. So kasi dba ang spx natin ay serum tapos yung mga instruments nag ssuck ng serum and they analyse it or cantify or determine the concentration of anaylte. Pwede raw na may masama na minute particles. Kapag merong silica yung gel. However, the resolution will be FILTER THE SERUM Memorize yong additive na present, inversion, para saan or the test. (Order of Draw- Syringe and Evacuated method) 7 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 In general, TANDAAN NIYO TO “MAGIC NUMBER WILL BE 8” kapag tinamad magreview para kabisaduhin ang lahat ng anticoagulant 8 is your magic number, siya ang piliin ninyo. Except wag niyo kakalimutan si light blue kasi 3-4 inversions siya. IN GENERAL, PAG WALA NA KAYONG ALAM, YOUR MAGIC NUMBER WILL BE 8 WITH A FEW EXCEPTIONS LIKE YOUR LIGHT BLUE AND SOME OTHER TUBES CONTAINING CLOT ACTIVATOR. PAG MERONG CLOT ACTIVATOR YAN, IN GENERAL, 5 IS YOUR MAGIC NUMBER. IN GENERAL PAG ANTICOAGULANT 8 IS YOUR MAGIC NUMBER, PAG MAY CLOT ACTIVATOR 5 IS YOUR MAGIC NUMBER. Although may exception pa rin naman don Table: Summary of Anticoagulants Used in the Order of Draw (Syringe and Evacuated Tube Method) STOPPER COLOR ADDITIVE COMMENTS YELLOW 8 Sodium Aids in bacterial recovery by Polyanethol Sulfonate inhibiting complement, phagocytes and (SPS) certain antibiotics ***As anticoagulant, chelates Calcium ★ yellow top tube the additive is SODIUM POLYANETHOL SULFONATE (SPS) IT IS BOTH ANTICOAGULANT AND ADDITIVE kasi SPS is already an anticoagulant that chelates calcium parang si citrate, EDTA, and oxalate. At the same time, aside from being anticoagulant it is also an anticomplement and antiphagocytic. It aids in bacterial recovery by inhibiting complement phagocytes and certain antibiotics. ★ Para saan ba kasi ang yellow top? For bacterial culture. If you want to take care of or recover bateria, of course you need to inhibit yung mga nag i-inhibit dun like complement, phagocyte and antibiotics. Yan pa ang isang role ni SPS aside from being anticoagulant. ★ Bakit number si yellow sa order of draw? Syempre bacteria, in order to minimize contamination, kapag hinuli mo siya or ginitna mo siya, na contaminate na. Imbis na sana yong bacteria lang na nacollect mo nandoon sa dugo, baka meron pang humalo from outside source bakac ma-ontaminate siya. So to minimize that, number one or una sa listahan ang yellow top. Light Blue 3-4 Sodium Citrate Chelates calcium For coagulation studies ★ Next si light blue, 3-4 inversions. The additive is sodium citrate, it chelates calcium studies like PT and APTT and platelet function assays. Pangalawa naman siya kasi pag pinatagal mo pa or sa gitna pa or sa dulo pa, naactivate na yong coagulation, anong manyayari? Falsely decrease or shorten ang PT and APTT mo kasi pag pinatagal mo ng pinatagal, hindi siya prinariotize mo over the other top, inuna mo yong ibang top. That's why the next priority is coagulation, para yung leeway niya na pag activate ng coagulation cascade, at least hindi masyadong , matagal Red glass; none Glass: None For chemistry Plastic: 5 Plastic: With Clot activator ★ Next is the red, in the form of glass or plastics. NO inversion ang glass kasi wala siyang additives. The glass itself is already a clot activator kasi may tinatawag tayong contact group, naalala niyo yon? Fibrinogen group, prothrombin group, etc. kaya sila tinawag na contact group kasi kapag dumikit sila sa glass naactive sila or sa collagen in addition to glass na activate sila, as such clot activator mo na ang glass. ★ Next is the plastic, unfortunately, ang plastic ay hindi clot activator that’s why meron kang clot activator. Dapat alam mo kung saan section ang request ng doctor sayo. Green 8 Sodium Heparin Inhibits Lithium Heparin Thrombin formation Enhances Antithrombin Ill - Next is the green, sodium heparin dapat hindi mo siya minemeasure (?? hindi maintindihan yong audio, my bad). It inhibits thrombin formation by enhancing or accelerating the reaction of antithrombin in facilitating activation of thrombin and anticoagulant. 8 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 Lavender/ Pink/ 8 Lavender Glass: Chelates Purple ★ K3EDTA Calcium For Lavender Plastic: most Hematology tests ★ K2EDTA Pink top: Spray-dried ★ KZEDTA - Next is the purple/pink/lavender top, 8 inversion. Chelates calcium and for hematology test because they also preserve the morphology. Gray 8 NaF Inhibits glycolysis Potassium Oxalate For Lithium iodoacetate Glucose Next, gray top. Sodium fluoride and lithium iodoacetate, sila ang additive but not anticoagulant. The anticoagulant present there is the oxalate or EDTA. Sodium fluoride can actually inhibit glycolysis for up to 3 days but is neither adequate nor effective in preserving glucose concentration, but as long as 3 days pero hindi naman ganyan umaabot yong pagrelease mo ng result sa glucose, kaya okay lang yong red top kasi hindi naman siya ganun katagal yong testing ng glucose. SIDENOTES: ★ This order of draw has been revised to reflect the increased use of plastic evacuated collection tubes. Plastic serum tubes containing a clot activator may cause interference in coagulation testing. Some facilities may continue using glass serum tubes without a clot activator as a waste tube before collecting special coagulation assays reflects change in CLSI recommended Order of Draw (Rodak) Noon kasi glass ang ginagamit pero according to OSHA ni recommend ang paggamit ng plastic however kasi plastic serum tubes containing clot activator,may cause interference in coagulation testing. Kaya ganyan ang nagging order of draw din. ★ Many laboratories have converted from glass to plastic collection tubes to minimize exposure to biohazardous material (e.g., blood) and broken glass; to lower biohazard waste disposal costs; and to comply with Occupational Safety and Health Administration (OSHA) guidelines mandating substitution. This change from glass to plastic has required a modification in the order of draw. Glass or plastic tubes with additives, including gel tubes, are drawn after the citrate tube (blue top) to avoid interference with coagulation measurements. Glass or plastic serum tubes, without a clot activator or gel separator, may be drawn before the coagulation tubes are drawn consistent with CLSI. (Henry) Dahil dito nabuo yong order of draw (explanation yan nasa taas teh) (May sinabi si Sir kaso hindi maintindihan yong audio, nagstop parang robot) Medj choppy na po Doc, choppy na po talaga Doc. Kaya na pansin niyo, yung glass na red ay after ng blue top. Yung red na merong gel or clot activator after din ng blue top. Kaya sa order of draw, PRIORITY si CITRATE, except kapag may culture ka. Glass or plastic serum tubes, without a clot activator or gel separator, may be drawn before the coagulation tubes are drawn consistent with CLSI. Kaya pwede yong yellow top na mauna kasya kay light blue top kasi hindi naman siya necessarily na, wala naman siyang clot activator or gel separator. At tsaka to prevent or to minimize contamination. 9 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 ★ If a winged blood collection set (BUTTERFLY) for venipuncture (Yong winged or butterfly, kadalasan siyang ginagamit sa wrist. Although kaya pa naman nating mag phlebotomy using syringe sa may wrist) ★ Paano naman order of draw kapag winged blood collection set? and a coagulation (citrate) tube is the first specimen tube to be drawn, a discard tube should be drawn first. To ensure a proper blood to citrate ratio, use the discard tube to fill the air space with blood. The discard tube does not need to be completely filled. The coagulation (citrate) tube is the first specimen tube to be drawn, a discard tube should be drawn first. Wag ka agad magcocollect sa light blue top or wag mong gagamitin yong first collection mo kasi dapat discard mo muna yon. Not necessarily na ifill up the whole discard tube, usually ang discard tube natin is light blue top, basta ang mahalaga natanggal mo yong hangin, diba nakakita na kayo ng butterfly or winged blood collection set? Diba meron siyang maliit na tubo, yong tube doon na dinasaluyan ng blood papunta sa top or stopper (may sinabi pa siya kaso nagchoppy ulit) So need mong tanggalin muna yong hangin, doon sa loob ng tubo para maasure na natanggal yong hangin meron kang discard tube ○ The waste tube must also be a light-blue top tube or a tube that contains no additives. This waste tube is drawn first to remove the air in the tubing of the winged collection device. Once blood flows through the tubing, the waste tube can be removed and discarded. The waste tube does not need to be filled (Hindi kailangan punoin yung discard tube. As long as natanggal mo yung hangin sa tubing okay na yun.) ○ If the air is not displaced from the tubing into a waste tube, it will be drawn into the tube used for testing and cause a short-fill of the tube (If hindi mo tinanggal yung hangin ito mangyayare).. ○ At dahil diyan Less blood volume in the tube alters the blood-to-anticoagulant ratio needed for coagulation studies (labce.com) result niya is falsely prolong kasi dumami yong anticoagulant mo kasi short fill yong tube mo. table: summary of other anticoagulants If order of draw is not the topic, paano naman yung ibang toppers na hindi kasali sa order of draw. Or not necessarily kasali sa order of draw or usually dahil kasi hindi siya common na ginagawa. STOPPER/COLOR ADDITIVE COMMENTS Yellow Acid Citrate Dextrose RBC and WBC preservative For HLA phenotyping and paternity testing The yellow top, aside from it containing the (SPS) sodium polyanethol sulfonate nag lalaman din siya ng Acid citrate dextrose. Ano yung anticoag dito? CITRATE – Ano si dextrose? sugar. Ang sugar or carbohydrates ay source of energy for RBC and WBC, kaya ginagamit siya for BB. Sa BB if nag ccollect ng blood units, ang laman niya ay ACID CITRATE DEXTROSE. Although, yellow top, kapag mag pperform ka ng HLA phenotyping and paternity testing (discuss sa IS/BB) Yellow top na nag nalalaman ng acid citrate dextrose Black Sodium Citrate Chelates Calcium For ESR Black top on the other hand contains Sodium Citrate, 3.2% at a ratio of blood to anticoag, 4:1 Kung black top, tatanungin ni sir sa order of draw, in the order of draw, saan or kailan ka mag ccollect using the black top? Might as well after light blue top or after yellow top kapag walang test sa coagulation test Gold, Red, Gray Orange, Royal Clot Activator Accelerates clotting For STAT Blue chemistries Have clot activator that accelerates clotting (discuss daw ni sir nung MTLBE) FOR STAT CHEMISTRY. Compared to an ASAP spx. which will you prioritise? The STAT SPX more than the ASAP spx. BEcause according to Henry, if a test is labeled as a STAT spx, Highest priority will be given to it. Pero hindi ini-STAT yung culture kasi may incubation time needed at need nila time to grow. Bakit STAT chemistry? Kasi meron siyang clot activator na nag a-accelerate ng clotting. From 60mins na standard down to 30mins. or as fast as 5mins. Kapag ang clot activator ang ginamit mo ay Thrombin, from 60mins ma accelerate siya magiging 5mins nalang Kaya kapag nag mamadali or kailangan e-prioritize yung patient (Choppy sir), PARTICULARLY your ORANGE, Although meron din si Royal blue. BTW si Royal blue ay walang separator gel, meron lang siyang Clot activator ( Si GORG or si G OR G) yun yung mga may both separation gel and clot activator. PERO SI ROYAL BLUE, may clot activator pero walang separator or thixotropic gel 10 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 Tan Glass: Sodium Heparin Sodium Heparin inhibits thrombin Plastic: K2EDTA K2EDTA chelates calcium Both of which are used in Lead Testing White EDTA with gel Chelates calcium For molecular diagnostics Chelates calcium for advance studies, for example PCR, branch DNA testing, or molecular assays/diagnostics Got from henry, ano naman ang effect ng anticoagulants and additives on blood test. 1. EDTA a. Bakit hindi ginagamit sa Chemistry? (yung sa table na Alkaline phosphate hanggang Calcium Iron) Pang chemistry, syempre if chemistry red top dba. Para saan ba ang EDTA? Sa hema. Kaya if nakita mo na hindi pang hema ekis na. b. Bakit decrease ang effect ni calcium and iron? kasi ang MOA niya ay, callates or inhibits calcium, pt/aptt. Although hematology siya, may tendency kasi na mag increase or falsely prolonged ang PT and APTT, as such light blue top c. Sodium and potassium din, kasi ito syempre pwedeng heparin or red top kasi falsely increase ang effect niya for sodium and potassium 11 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 2. OXALATE a. (CHOPPY) pero like your c hemolysis test??? Rightt?? b. Bakit yung calcium decrease? Kasi ano ba ang MOA ng oxalate (kalase?) calcium,forming insoluble complex c. Cell morphology is distorts in peripheral blood film d. Calcium, decreases din. Ano ba ang MOA ng citrate, calates or binds with calcium forming soluble complexes e. Sodium potassium, also the effect is increased. that's why heparinized, blood plasma, or non-anticoagulated tube f. Labile coagulation factors, ano yung dalawang yun? FACTORS V and FACTORS VIII, yan yung pinepreserve niya kaya anticoagulant of choice siya for PT/APTT 3. HEPARIN a. T3 and T4 – increase kaya red top pa rin b. The most important – WRIGHT’S STAIN i. It causes blue background or gray background kasi as exemplified by rodak. Kaya hindi mo pwedeng gamitin ang heparin to make PBS because it cause blue background in wright stain. Common sense na no need for explanation 4. FLUORIDES a. Cell morphology distorted din parang si Oxalate. b. Para saan si fluoride? Glycolysis, wala siyang pakialam sa mga enzyme na nasa table. More on ted top sila Blood collection tubes with Hemogard M (BD) closure protect you from blood which might splatter when the tube is opened. The rubber stopper is recessed inside the plastic shield, preventing exposure to blood present on the stopper. -Labce.com Iba na ang stopper na ginagamit ng Lorma, kasi modern na tayo. Sa pic, yan na yung na kikita natin - A plastic shield or yung tinatawag nating hemogard closure. Yung mga conventional kasi na mga stopper, yung stopper presented sa pic, may rubber sa loob to prevent splatter. SYmpre bubuksan si stopper dba, at least yung stopper mo hindi pinapasok sa loob mismo ng tube, may tendency kasi for vacuum, pwedeng mag splatter. - Your blood collection tube w/ Hemogard M (BD) closure protect you from blood which might splatter when the tube is opened. 12 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 13 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL  BATCH SORBETES ‘25 Aralin yung pic yung gold top, red or gray top, merong gel for serum separation. Chemistry sila. If may clot activator na mminimize siya from 60mins down to 30mins Ano role ng inversion? Ensure mixing of additive to the blood Anong rule natin? Kapag merong separator gel or clot activator your magic number is 5, except for royal blue. for royal blue kasi, 8. ALL ANTICOAGULANTS WILL BE 8, except citrate and of course except those with additives clot activator or serum separtor gel. The rest will be 8 Sa mga tinatamad, 8 ang isasagot kapag hindi nag review Light or green or gray, nag lalaman sila ng heparin and gel plasma preparation. For Chemistry, particularly for potassium, sodium and pwede ring ABG pero since may heparin na mismo yung syringe na ginagamit for ABG kaya hindi na sila kadalasan ginagamit, na kailangna mong ilagay dun sa tube for ABG analysis or for those test na gumagamit ng heparin. Kasi meron yung mga syringe na nag lalaman na ng heparin sa mismong barrel niya RED – Pag glass, no clot activator kasi the glass itself is already clot activator. For plastic meron, again, for clot activator or clot additive, 5. ○ 60mins for glass ○ Pero pag may clot activator down to 30mins Now if your clot activator is thrombin, napaka bilis, 5mins For orange, huwag kakalimutan, STAT. Kasi may clot activator, 5mins Royal blue – CHOPPYY si sir – yung isang plastic meron K2EDTA GRAY – Meron ding gray top na nag lalaman ng sodium fluoride lang (ARGHH, CHOPPY ULET, PANOORIN NIYO NALANG RECORDED LEC) TAN – K2EDTA for lead determination Yellow – blood culture. ANo ginagawa niya aside from (collating?) with calcium? Antibacterial, anticomplement, and antiphagocytic para marecover yung bacteria ○ ACD is for BB, particulary HLA phenotyping and paternity testing White nag lalama nng EDTA ○ HIGHLIGHT LANG SI WHITE TOP, for advance molecular diagnostic test like PCR Light blue top nag lalaman ng sodium citrate, Citrate, Theophylline, Adenosine, Dipyridamole (CTAD) ○ Anticoag sa CTAD ay si CITRATE, si Theophylline at Dipyridamole ay antiplatelet sila. So if you’re studying coagulation the light blue talaga ○ Say for example, according to studies, can be considered daw ang as a universal of the coagulation. But the term universal, isa daw itong anticoagulant na pwede mong gamitin universal, hindi lang sa coagulation, pwede rin sa chem, pwede sa CBC. Kaya may potential daw siyang to be used as a universal anticoagulant para hindi ka malito anong topper ang gagamitin mo (NAG CHOPPY NA YUNG IBA) 14 | LIM, JULIANA NICOLE B.; TORES, ANGELICA; LAYMAN, CRYSTAL